Responsive microgel and methods related thereto

ABSTRACT

A responsive microgel is provided which responds volumetrically and reversibly to a change in one or more aqueous conditions selected from the group consisting of (temperature, pH, and ionic conditions) comprised of an ionizable network of covalently cross-linked homopolymeric ionizable monomers wherein the ionizable network is covalently attached to an amphiphilic copolymer to form a plurality of ‘dangling chains’ and wherein the ‘dangling chains’ of amphiphilic copolymer form immobile micelle-like aggregates in aqueous solution. A responsive microgel is further provided that comprises at least one therapeutic entity and delivers a substantially linear and sustained release of the therapeutic entity under physiological conditions.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a divisional of U.S. patent application Ser. No.10/298,808 filed Nov. 18, 2002 which claims the benefit of U.S.Provisional Application Ser. No. 60/352,200 filed on Jan. 29, 2002 allof which are hereby incorporated in their entirety by reference to thisapplication.

FIELD OF THE INVENTION

The present invention relates to microgels comprised of an ionizablenetwork covalently attached to an amphiphilic copolymer, which formsaggregates capable of solubilizing drugs in aqueous solution. Theresponsive microgel reversibly responds volumetrically to factors suchas temperature, pH, and ionic conditions. Particularly, the responsivemicrogel is able to imbibe or solubilize a large amount of therapeuticagent and deliver a substantially linear and sustained release oftherapeutic agent under physiological conditions.

BACKGROUND OF THE INVENTION

Volumetric changes (shrinking or swelling) of temperature-sensitivemicrogel particles dispersed in water is an intraparticle phenomenon,but it is known in the art that also interparticle aggregation takesplace during the collapse transition. Very stable polymer dispersionshave been synthesized, for instance, using poly(ethylene oxide), PEO, asa stabilizing agent. Ottewill, R. H., et al., Colloid Polym. Sci. 1995,273, 379.

The most widely studied class of responsive polymers are temperatureresponsive poly(alkylacrylamides), specificallypoly(N-isopropylacrylamide). Shibayama, M., et al., Advances in PolymerScience; Springer-Verlag: Berlin, 1993; 109, pp 1-62; Pelton, R. Adv.Colloid interface Sci. 2000, 85, 1-33. However, poly(alkylacrylamides)are perceived to be toxic, especially in biomedical applications. L. E.Bromberg and E. S. Ron, Adv. Drug Delivery Revs., 1998, 31, 197-221.Furthermore, nonionic nature of poly(alkylacrylamides) prevents creatingion-sensitive microgels.

Synthesis of polymers comprised of polyethers such as poly(ethyleneoxide), poly(propylene oxide) and their copolymers grafted ontopoly(acrylic acid) and other polyelectrolytes such aspoly(2-acrylamido-2-methylpropanesulfonic acid), polyethyleneimine andthe like are known in the art. See, e.g., Hourdet, D., et al., Polymer(1994), 35(12), 2624-30; L'Alloret, et al., Colloid Polym. Sci. (1995),273(12), 1163-73; L'alloret, F.; Maroy, P., et al., Revue de l'institutFrancais du Petrole (1997), 52(2), 117-128; Hourdet, D. et al.,Macromolecules; 1998; 31(16); 5323-5335; Schiumberger, D. C., EPOPublication 0 583 814 A1, 1993; 0 629 649 A1, 1994; Hoffman, et al.,Advanced Biomaterials in Biomedical Engineering and Drug DeliverySystems, Ogata, N., et al., Kim, S. W., Feijen, J., Okano, T., Eds.,Springer: Tokyo, 1996; pp 62-66; Hoffman, A. S., et al., Proc. Inte.Symp. Controlled Release Bioact. Mater. (1995), 22, 159; Chen, G., etal., Proc. Int. Symp. Controlled Release Bioact. Mater. (1995), 22, 167;Hoffman, A. S.; E. S. Ron, L. E. Bromberg, M. Temchenko, End ModifiedThermal Responsive Microgels, U.S. Pat. No. 6,316,011.

These polymers are synthesized by conversion of one or both terminalOH-groups of a polyether into a more active group such as NH₂—, SH—,followed by grafting of the resulting modified polyether onto thebackbone of a chosen polyelectrolyte. Structures that could result fromthese syntheses may comprise, for example, un-cross-linked PLURONIC®copolymer bonded to poly(acrylic acid). See, e.g., A. S. Hoffman, etal., Advanced Biomaterials in Biomedical Engineering and Drug DeliverySystems, N. Ogata, S. W. Kim, J. Feijen, T. Okano, eds., Springer,Tokyo, 1996, pp. 62-66; A S Hoffman, et al., Polym. Prepr., 38: 524-525,(1997); G. Chen, et al., Poly(ethylene glycol) Chemistry and BiologicalApplications, edited by J. Milton Harris, S. Zalipsky, eds., AmericanChemical Society, Washington, D.C., (1997), ACS Symposium Series 680,Chapter 27, pp. 441-457; A. S. Hoffman, et al., Frontiers in BiomedicalPolymer Applications, edited by R. M. Ottenbrite, Technomic PublishingCo., Lancaster, Pa. 1999, Vol. 2, pp. 17-29; A. S. Hoffman, et al.,Block and graft copolymers and methods related thereto, Int. Pat. Appl.WO 95/24430.

Alternatively, syntheses known in the art can result in chemicallycross-linked networks (gels, microgels, or nanogels) if both termini ofthe polyether are chemically modified. See, e.g., U.S. Pat. No.6,316,011; L. Bromberg, Crosslinked poly(ethylene glycol) networks asreservoirs for protein delivery, J. Appl. Polym. Sci., 59(1996)459-466;L. Bromberg, Temperature-sensitive star-branched poly(ethyleneoxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) networks, Polymer,39(23)(1998) 5663-5669; Law, T. K., et al., Int. J. Pharm., 1984, 21,277; Ping, Q.; Law, T. K., et al., Int. J. Pharm., 1990, 61, 79.

However, chemical moieties (amide or other) required to accomplishlinkage between an amphiphilic copolymer such as polyether and thepolyelectrolyte (i.e. a chemical group absent in parent polyether andpolyelectrolyte) are generally toxic and unacceptable for use inpharmaceutical and other applications. Copolymers that comprisechemically unmodified amphiphilic copolymer and polyelectrolyte bondedonly through carbon-carbon bond, where such toxicity issues are avoided,are known in the art. See, e.g., L Bromberg, et al., Responsive polymernetworks and methods of their use, U.S. Pat. No. 5,939,485; E. S. Ron,et al., Compositions for pharmaceutical applications, Int. Patent Appl.WO 98/06438; E. S. Ron, et al., T. H. E. Mendum, Compositions forcosmetic applications, Int. Patent Appl. WO 98/50005; L Bromberg,Hydrophobically modified polyelectrolytes and polyelectrolyteblock-copolymers, Handbook of Surfaces and Interfaces of Materials, H.S. Nalwa, ed., Academic Press, 2001, Vol. 4, Chapter 7; L Bromberg,Biomedical applications of hydrophobically modified polyelectrolytes andpolyelectrolyte block-copolymers, S. Tripathy, J. Kumar, H. S. Nalwa,eds. Handbook of Polyelectrolytes and Their Applications. AmericanScientific Publishers, Stevenson Ranch, Calif., 2002, Vol. 1, Chapter51; L. E. Bromberg, T. H. E. Mendum, M. Orkisz, E. S. Ron, E. C. Lupton,Applications ofpoly(oxyethylene-b-oxypropylene-b-oxyethylene)-g-poly(acrylic acid)polymers (Smart Responsive microgel™) in drug delivery, Proc. Polym.Mater. Sci. Eng., 76: 273-275, 1997; M. J. Orkisz, et al., Rheologicalproperties of reverse thermogelling poly(acrylicacid)-g-poly(oxyethylene-b-oxypropylene-b-oxyethylene) polymers (SmartResponsive microgel™), Proc. Polym. Mater. Sci. Eng., 76: 276-277, 1997;L. Bromberg, et al., Interpenetrating networks of Poloxamer copolymersand poly(acrylic acid) as vehicles in controlled drug delivery, J.Control. Release, 48 (2, 3): 305-308, 1997; L. E. Bromberg, T. H. E.Mendum, M. J. Orkisz, E. C. Lupton, E. S. Ron,Polyoxyethylene-b-polyoxypropylene-b-polyoxyethylene-g-poly(acrylicacid) polymers (Smart Responsive microgel™) as a carrier in controlleddelivery of proteins and peptides, Polym. Prepr., 38(2): 602-603, 1997;L. E. Bromberg, et al., Bioadhesive properties ofpolyoxyethylene-b-polyoxypropylene-b-polyoxyethylene-g-poly(acrylicacid) polymers (Smart Responsive microgel™), Polym. Prepr., 38(2):626-627, 1997; L. Bromberg, A novel family of thermogelling materialsvia C—C bonding between poly(acrylic acid) and poly(ethyleneoxide)-b-poly(propylene oxide)-b-poly(ethylene oxide), J. Phys. Chem. B,102: 1956-1963, 1998; L. E. Bromberg, E. S. Ron, Protein and peptiderelease from temperature-responsive gels and thermogelling polymermatrices, Adv. Drug Delivery Revs., 31:197-221, 1998; L. E. Bromberg, M.G. Goldfeld, Self-assembly in aqueous solutions of hydrophobicallymodified poly(acrylic acid), Polym. Prepr., 39(2):681-682, 1998; L.Bromberg, Scaling of rheological properties of responsive microgels fromassociating polymers, Macromolecules, 31: 6148-6156, 1998; L. Bromberg,Self-assembly in aqueous solutions of polyether-modified poly(acrylicacid), Langmuir, 14: 5806-5812, 1998; L. Bromberg, Polyether-modifiedpoly(acrylic acid): synthesis and properties, Ind. Eng. Chem. Res., 37:4267-4274, 1998; L. Bromberg, Properties of aqueous solutions and gelsof poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethyleneoxide)-g-poly(acrylic acid), J. Phys. Chem B, 102: 10736-10744, 1998; L.Bromberg, L. Salvati, Bioactive surfaces via immobilization ofself-assembling polymers onto hydrophobic materials, Bioconjugate Chem.,10: 678-686, 1999; L. E. Bromberg, Interactions between hydrophobicallymodified polyelectrolytes and mucin, Polym. Prepr., 40(2): 616-617,1999; L. E. Bromberg, D. P. Barr, Aggregation phenomena in aqueoussolutions of hydrophobically modified polyelectrolytes. Macromolecules,32: 3649-3657, 1999; P. D. T. Huibers, et al., Reversible gelation insemidilute aqueous solutions of associative polymers: a small-angleneutron scattering study, Macromolecules, 32: 4889-4894, 1999; L.Bromberg, E. Magner, Release of hydrophobic compounds from micellarsolutions of hydrophobically modified polyelectrolytes, Langmuir, 15:6792-6798, 1999; L. Bromberg, M. Temchenko, Loading of hydrophobiccompounds into micellar solutions of hydrophobically modifiedpolyelectrolytes, Langmuir, 15: 8627-8632, 1999; L. E. Bromberg, M.Temchenko, R. H. Colby, Interactions among hydrophobically modifiedpolyelectrolytes and surfactants of the same charge, Langmuir, 16:2609-2614, 2000; N. Plucktaveesak, et al., Effect of surfactants ongelation threshold temperature in aqueous solutions of hydrophobicallymodified polyelectrolyte, Proc. XIIIth International Congress onRheology, Cambridge, UK, 2000, Vol. 3, pp. 307-309; L. Bromberg,Enhanced nasal retention of hydrophobically modified polyelectrolytes,J. Pharm. Pharmacol., 53: 109-114, 2001; L. Bromberg, Interactions amongproteins and hydrophobically modified polyelectrolytes, J. Pharm.Pharmacol., 53: 541-547, 2001; A. K. Ho, L. E. Bromberg, et al., Solutediffusion in solutions of associative polymers, Langmuir, 17: 3538-3544,2001; R. H. Colby, et al., Critical incorporation concentration ofsurfactants added to micellar solutions of hydrophobically modifiedpolyelectrolytes of the same charge, Langmuir, 17: 2937-2941, 2001; L.Bromberg, Synthesis and self-assembly of poly(ethyleneoxide)-b-poly(propylene oxide)-b-poly(ethylene oxide)-g-poly(acrylicacid) gels, Ind. Eng. Chem. Res., 40: 2437-2444, 2001; L. Olivieri, etal., Study of the breakup under shear of a new thermally reversibleW/O/W multiple emulsion, Pharm. Res., 18: 689-693, 2001; A. K. Ho, etal., Hydrophobic domains in thermogelling solutions ofpolyether-modified poly(acrylic acid), Langmuir, 18: 3005-3013, 2002.

One-step methods of copolymer synthesis are known in the art, e.g., L.Bromberg, J. Phys. Chem. B, 1998, 102, 1956-1963; L. Bromberg, Ind. Eng.Chem. Res., 1998, 37, 4267-4274; P. D. T. Huibers, et al.,Macromolecules, 1999, 32, 4889-4894. However, these prior art graft-combcopolymers are not permanently cross-linked and therefore cannot respondvolumetrically to changes in their environment.

Previous Work

A variety of formulation approaches have been developed aiming at theenhancement of the drug residence time and to lowering the release rate,while maintaining the mucoadhesive properties of the polyelectrolyte.See, e.g., K. J. Himmelstein, et al., U.S. Pat. No. 5,599,534; T XViegas, et al., U.S. Pat. No. 5,292,516; R Joshi, et al., Pharm DevTechnol. 4: 515-522 (1999); Joshi, et al., U.S. Pat. No. 5,252,318.Typically, a polyelectrolyte is mixed with more hydrophobic polymer toresult in a blend with enhanced drug-polymer interactions and higherviscosity. It is preferred, however, that a liquid drug-polymerformulation gel at the site of administration. Such in situ gellingsystems undergo reversible sol-gel transitions in response totemperature, pH, or ion composition of the fluids. However, physicalblends are colloidally unstable and either phase separate or dissociateat physiological pH. See, e.g., L Bromberg, Handbook of Surfaces andInterfaces of Materials, H. S, Nalwa, ed., Academic Press, 2001, Vol. 4,Chapter 7. Therefore, these blends fail to provide a linear, sustainedrelease of a hydrophobic or amphiphilic compound such as imbibed orloaded drug, for example, in drug delivery applications.

Previous structures that result from the linking of the amphiphiliccopolymers and polyelectrolyte though carbon-carbon bond (See FIG. 1,structure I (A.) and FIG. 2 structure III, for example) can formphysical gels in water due to aggregation of the hydrophobic segments ofthe amphiphilic copolymers at certain temperatures and concentrations.However, by definition, previous gels are unstable upon dilution due todissociation of the physical aggregates below a certain concentration.Accordingly, due to dissociation under physiological conditions,previous gels were not able to provide a linear, sustained release of ahydrophobic or amphiphilic compound such as imbibed or loaded drug, forexample, in drug delivery applications. Further, previous structuresthat result from chemical linking on both termini (See FIG. 1, structureII (B.), for example) can form stable, chemically cross-linked networks.However, due to the steric constrains imposed by chemical linking onboth termini, the hydrophobic parts of the amphiphilic copolymer areunable to aggregate at well-defined temperatures and concentrations.Therefore, nanosized aggregates do not form within the gel network. As aresult, such previous gels also were not able to provide a linear,sustained release of a hydrophobic or amphiphilic compound such asimbibed or loaded drug, for example, in drug delivery applications.

SUMMARY OF THE INVENTION

A responsive microgel is provided which responds volumetrically andreversibly to a change in one or more aqueous conditions selected fromthe group consisting of (temperature, pH, and ionic conditions)comprised of an ionizable network of covalently cross-linkedhomopolymeric ionizable monomers wherein the ionizable network iscovalently attached to an amphiphilic copolymer to form a plurality of‘dangling chains’ and wherein the ‘dangling chains’ of amphiphiliccopolymer form immobile micelle-like aggregates in aqueous solution.

A responsive microgel is further provided that comprises at least onetherapeutic entity and delivers a substantially linear and sustainedrelease of the therapeutic entity under physiological conditions.

A responsive microgel is also provided wherein the ionizable network ofcovalently cross-linked homopolymeric ionizable monomers is selectedfrom the group consisting essentially of (poly(acrylic acid),poly(methcarylic acid), poly(4-vinylpyridinium alkyl halide),poly(sodium acrylate), poly(sodium methacrylate), sulfonatedpolyisoprene, and sulfonated polystyrene).

A further responsive microgel is provided wherein an amphiphiliccopolymer is comprised of (poly(ethylene oxide) and a monomer selectedfrom the group consisting essentially of (poly(propylene oxide),poly(butylene oxide), polystyrene, polyisobutylene, poly(methylmethacrylate), and poly(tert-butyl acrylate)).

A method of making a responsive microgel is also provided comprising:

a) providing, an ionizable monomer, a divinyl cross-linker, a freeradical, and a amphiphilic copolymer; and

b) copolymerizing the ionizable monomer with the divinyl cross-linker toproduce an ionizable network, while

c) abstracting hydrogen from the amphiphilic copolymer with the freeradical to progress a chain transfer reaction wherein the amphiphiliccopolymer is covalently bonded onto the ionizable network to produce theresponsive microgel.

A method of administering an effective amount of at least onetherapeutic entity to a patient is further provided which comprisesadministering a responsive microgel comprising an effective amount of atleast one therapeutic entity.

A method is provided for administering at least one therapeutic entityto a patient which entity is selected from the group consisting ofsubstrates of ABC transporters such as P-glycoprotein, MRP1-MRP9; ABChalf-transporters such as BCRP; other transporters that are involvedinto a limited drug transport across small intestinal epythlium;cerebral endothelium and other barrier tissues in the body, as well assubstrates of metabolic enzyme isoforms without limitation, cytochromeP-450; esterase; epoxide hydrolase; alcohol dehydrogenase; aldehydedehydroganase; dihydropyrimidine dehydroganase; NADPH-quinoneoxidoreductase; uridine 5′-triphosphat glucoronosyltransferase;sulfotransferase; glutatione S-transferase; N-acetiltransferase;histamine methyltransferase; catechol-o-methyl transferase; thiopurinemethyltransferase. This group of therapeutic agents include withoutlimitation doxorubicin and other anthracyclines, mitoxantrone, mitomycinC, metatrexate, paclitaxel, docetaxel and other taxanes, topotecan antother camptotecines, cysplatin, carboplatin, oxaliplatin and otherplatinum complexes; megesterol acetate and other steroids; carvediloland other beta-blocking agents; azidothymidine, fludarabine and othernucleoside containing agents in their dephospho, mono-, di- andtri-phosphorylated forms; vinblastine, vincristine and other vinkaalkaloids; etoposide and other podophilotoxins.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows previous structures (A.) (structure I) and (B.) (structureII) that result from the linking of amphiphilic copolymers andpolyelectrolyte though carbon-carbon bonds.

FIG. 2 shows another previous structure (structure III) that resultsfrom the linking of amphiphilic copolymers and polyelectrolyte thoughcarbon-carbon bonds.

FIG. 3 is a general macro-illustration of the overall ionizable networkand the covalently attached ‘dangling chains’ (amphiphilic copolymer)structure of the responsive microgel described herein.

FIG. 4 is a flowchart illustration of one-step synthesis of responsivemicrogels of the present invention.

FIG. 5 shows the kinetics of the release of doxorubicin and PLURONIC®L61 through dialysis membrane with and without a responsive microgeldescribed in Example I.

FIG. 6 shows the kinetics of the drug diffusion in the controlexperiment described in Example I.

FIG. 7 shows the kinetics of doxorubicin release from a responsivemicrogel.

FIG. 8 shows a very slow, sustained release of PLURONIC® L61 from anexample responsive microgel of the present invention.

FIG. 9 shows the results of equilibrium swelling experiments, e.g., highabsorbency as a function of the subchain length.

FIG. 10 shows equilibrium swelling of microgel particles in deionizedwater at 15° and 37° C. as a function of pH. Degree of cross-linking inmolar percent is indicated.

FIG. 11 shows equilibrium swelling of microgel particles in deionizedwater at pH 7.0 as a function of temperature. Degree of cross-linking inmolar percent is indicated.

FIG. 12 shows the effect of temperature on hydrophobic compounds inresponsive microgel suspension.

FIG. 13 shows the equilibrium uptake of doxorubicin by microgels of thepresent invention as a function of pH at 37° C.

FIG. 14 shows transepithelial transport of doxorubicin from thebasolateral to the apical (b→a) and from the apical to the basolateral(a→b) side of the Caco-2 monolayers. Initial concentration ofdoxorubicin in the donor compartment was 3 μM, and the doxorubicinconcentration in the receiver compartment was assayed by fluorescence(Fl, filled points) or HPLC (open points). Besides doxorubicin in thedonor compartment, no further additive was applied in DMEM. Thetransport was characterized by linear fits (R²>0.98 in all cases). Theapparent permeability obtained by the fluorescence and HPLC assay in theb→a tests was (2.81±0.03)×10⁻⁶ and (2.75±0.03)×10⁻⁶ cm/s, respectively,while the P_(a) values obtained in the a→b tests were (6.07±0.04)×10⁻⁷and (5.79±0.04)×10⁻⁷ cm/s using fluorescence and HPLC assays,respectively. Typically, the transport experiments did not exceed 2.5 hin duration.

FIG. 15 shows cumulative transport of ¹⁴C-mannitol (MW 182.2 Da) acrossCaco-2 cell monolayers. Data are expressed as mean ±S.D. of three tofive experiments.

FIG. 16 shows the effect of polymers (0.5 mg/mL each) on TEER of Caco-2cell monolayers. Inset shows TEER recovery of Caco-2 cell monolayersafter removal of the polymers. Data are expressed as mean ±S.D. of threeto five experiments.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to stable chemically cross-linked networks(gels) of a polyelectrolyte wherein ‘dangling chains’ of at least oneamphiphilic co-polymer are bonded thereto through carbon-carbon bonding.The dangling chains are capable of forming intra-network micelle-likeaggregates. The aggregates possess the ability to imbibe a largequantity of, for example, hydrophobic or amphiphilic compounds. Due tothe formation of mixed aggregates, the responsive-microgel networks ofthe present invention display linear and sustained release ofhydrophobic or amphiphilic compounds in aqueous milieu. Further, theformation of micelle-like aggregates within the chemically cross-linkedpolyelectrolyte network of the present invention is reversible.

The responsive microgel described herein is, for example, (1) able toimbibe large quantities of at least one ionic, amphiphilic, orhydrophobic compound, and (2) forms micelle-like aggregates within itsstructure when in aqueous solution and (3) allows for a sustained,substantially linear release of the compound in vitro and/or in vivo,for example, under the temperature, pH, and ionic composition ofphysiological conditions. A preferred embodiment of the presentinvention is method of delivering an effective amount of at least onetherapeutic agent to a patient comprising administering an effectiveamount of a responsive microgel of the present invention, whichcomprises at least one therapeutic agent. The responsive microgels ofthe present invention are suitable for oral administration, for example,and hence the oral delivery of therapeutic agents.

Drug release kinetics from example responsive microgels of the presentinvention are provided herein. Example I shows that loading ofcorresponding drugs into a responsive microgel greatly affected thekinetics of release. The drugs loaded into the microgel exhibited slow,sustained release kinetics. Kinetics of doxorubicin release from aresponsive microgel is shown in FIG. 7. Three cationic and one unchargeddrug was loaded onto the microgel in Example VII, all of which arecurrently in clinical use as anticancer drugs. Doxorubicin,mitoxantrone, and mitomycin C are mono-, di-, and trivalent cationicweak bases, respectively. Taxol is uncharged (hydrophobic). The abilityof responsive microgels of the invention to effectively load and holdtaxol, combined with mucoadhesive properties is a feature important fordelivery of taxol and other hydrophobic solutes such as steroidhormones. The taxol loading capacity provides additional evidence to themechanism of taxol solubilization into micelle-like aggregates withinthe responsive microgels. Drug loading via ion-exchange are illustratedusing the potent chemotherapeutic drug doxorubicin.

The responsive microgel of the present invention comprises tworesponsive components: An amphiphilic copolymer (nonionic copolymer)capable of aggregation in response to a change in temperature; and, anionizable, covalently cross-linked polymeric network of monomers whichresponds volumetrically to changes in aqueous conditions such as pH orionic composition by swelling or collapsing. Since both responsivecomponents, i.e., the nonionic copolymer and the cross-linkedpolymermeric network of monomers which contain ionizable groups arebound through covalent bonds, each polymer has a chemical or mechanicalinfluence over the swelling of the other polymeric component. Theresulting responsive microgel exhibits volumetric changes in response tovariations in pH as well as temperature. See, Examples III-V. Theseresponsive microgel graft-comb copolymers dissolve freely in aqueoussolutions and self-assemble in response to changes in conditions such aspH and temperature.

The microgel covalently cross-linked polymer network of the presentinvention is comprised of at least one amphiphilic copolymer covalentlyattached (preferably a carbon-carbon bond from a single terminal regionof each amphiphilic copolymer) to an ionizable network(polyelectrolyte). The amphiphilic copolymer forms the ‘dangling chain’component of the responsive microgel which forms micelle-like aggregateswithin the covalently cross-linked polymer network in aqueous solution.See, FIG. 3 (structure of the responsive microgel).

The term “responsive” in reference to the microgel of the presentinvention refers to reversible phase transition characteristics, e.g.,volumetric change, which result from exposure to a change in one or moreenvironmental factors under aqueous conditions, such as temperature, pH,and ionic conditions. The microgels operate as described herein withinthe temperature range of about −4° C. to about 55° C., preferably fromabout 0 to about 37° C. The microgels will be collapsed at pH 1-3 suchas in stomach and swollen at pH exceeding the pKa of their carboxylicgroups, i.e. at pH>4.5 (fully swollen at pH 7.4, for example). The gelis collapsed (swelling degree preferably not exceeding 50 v/v % of waterper polymer) at acidic pH such as in stomach, but fully swollen(swelling degree preferably exceeding 100-5000 v/v % of liquid perpolymer) in the intestine. The gels protect the therapeutic entity,e.g., embedded drug, and hold it without release when collapsed, butrapidly release when swollen. The range of operation of the microgels ofthe present invention are solutions of ionic strength preferably below 1M, or from 0M to 5 M, for example. A change in these environmentalfactor(s) affects the responsive microgel by causing the structure toundergo a reversible volumetric change in which the gel increases volumeby expanding (swelling) or decreases volume by collapsing (contraction).

Phase transitions in gels may be explained, for example, by thefollowing equation. One may determine the effect of ionic groups on thereduced chemical potential (Δμ₁) for solvent in an isotropically swollengel network: $\begin{matrix}{{\Delta\quad\mu_{1}} = {{\left( {\mu_{1} - \mu_{1}^{0}} \right)/{RT}} = {{\ln\left( {1 - v_{2}} \right)} + v_{2}}}} \\{{{f(\lambda)} + \frac{\Delta\quad\mu_{i}}{RT}} = {\ln\quad a_{i}}}\end{matrix} + {\chi\quad v_{2}^{2}} +$where a₁ is the activity of the solvent in the network, χ is theinteraction parameter, v₂ is the volume fraction of the polymer, f(λ) isthe function of the deformation tensor, Δμ_(i) is the contribution tothe total chemical potential by the presence of ionic groups on thechains.

Example I describes the favorable linear release of monomeric PLURONIC®from the microgels. It was discovered that PLURONIC® 161, for example,has exceptionally low release rate and sustained release for over 10days due to the formation of mixed micelles between added PLURONIC® 161and PLURONIC® covalently grafted to a poly(acrylic acid) network in theprocess of synthesis. Such mixed, immobile micelles can providethermodynamically stable environment for the PLURONIC® solute, makingits effective partition coefficient between micelles and water to bevery low. These results are unique and exceptionally well suited for theintended application of the novel microgels in drug delivery.

Compositions

1. Ionizable Network

The ionizable network is a covalently cross-linked homopolymeric networkof ionizable monomers. The monomers of the ionizable network eachcontain at least one ionizable group. The ionizable network respondsvolumetrically to changes in aqueous conditions such as pH or ioniccomposition by swelling or collapsing. Preferred embodiments of thispolyelectrolyte network (onto which the amphiphilic copolymer ‘danglingchains’ are attached via C—C bond to form the responsive microgel) arecomprised of a monomer selected from the group consisting essentially of(poly(acrylic acid), poly(methcarylic acid), poly(4-vinylpyridiniumalkyl halide), poly(sodium acrylate), poly(sodium methacrylate),sulfonated polyisoprene, and sulfonated polystyrene).

Preferred polyanion-forming compounds include poly(acrylic acid),poly(methcarylic acid), and poly(2-ethylacrylic acid); preferredpolycation-forming compounds include polyethyleneimine andpolyethylenepiperazine. The hydrophilic blocks recited infra, (i.e., A.Hydrophilic Monomers and Polymers), can also be used in the compositionsdescribed herein either as an element of the ionizable network(polyelectrolyte).

A. Polyanion Forming Compounds

Ionizable compounds for the ionizable network of the present inventionalso include, but are not limited to, polyanion-forming compounds suchas poly(acrylic acid), poly(methacrylic acid), poly(maleic acid),poly(styrenesulfonic acid), poly(itaconic acid), poly(vinyl sulfate),poly(vinylsulfonic acid), poly(vinyl phosphate), poly(acrylicacid-co-maleic acid), poly(styrenesulfonic acid-co-maleic acid),poly(ethylene-co-acrylic acid), poly(phosphoric acid), poly(silicicacid), hectorite, bentonite, alginic acid, pectic acid, kappa-, lambda-and iota-carrageenans, xanthan, gum arabic, dextran sulfate,carboxymethyldextran, carboxymethylcellulose, cellulose sulfate,cellulose xanthogenate, starch sulfate and starch phosphate,lignosulfonates, karaya gum; polygalacturonic acid, polyglucuronic acid,polyguluronic acid, polymannuronic acid and copolymers thereof;chondroitin sulfate, heparin, heparan sulfate, hyaluronic acid, dermatansulfate, keratan sulfate; poly-(L)-glutamic acid, poly-(L)-asparticacid, deoxyribonucleic acid, ribonucleic acid, acidic gelatins(A-gelatins); starch, amylose, amylopectin, cellulose, guar, gum arabic,karaya gum, guar gum, pullulan, xanthan, dextran, curdlan, gellan,carubin, agarose, as well as chitin and chitosan derivatives having thefollowing functional groups in various degrees of substitution:carboxymethyl and carboxyethyl, carboxypropyl, 2-carboxyvinyl,2-hydroxy-3-carboxypropyl, 1,3-dicarboxylsopropyl, sulfomethyl,2-sulfoethyl, 3-sulfopropyl, 4-sulfobutyl, 5-sulfopentyl,2-hydroxy-3-sulfopropyl, 2,2-disulfoethyl, 2-carboxy-2-sulfoethyl,maleate, succinate, phthalate, glutarate, aromatic and aliphaticdicarboxylates, xanthogenate, sulfate, phosphate, 2,3-dicarboxy,N,N-di(phosphatomethyl)aminoethyl, N-alkyl-N-phosphatomethylaminoethyl.These derivatives may additionally comprise nonionic functional groupsin various degrees of substitution, such as methyl, ethyl, propyl,isopropyl, 2-hydroxyethyl, 2-hydroxypropyl and 2-hydroxybutyl groups,for example, as well as esters with aliphatic carboxylic acids, e.g.,(C₂ to C₁₈).

B. Polycation Forming Compounds

Examples of polycation-forming compounds for the ionizable network ofthe present invention also include, but are not limited to,poly(alkylenimines), especially poly(ethylenimine),poly-(4-vinylpyridine), poly(2-vinylpyridine),poly(2-methyl-5-vinylpyridine), poly(4-vinyl-N—C₁-C₁₈-alkylpyridiniumsalt), poly(2-vinyl-N—C₁-C₁₈-alkylpyridinium salt), polyallylamine,polyvinylamine, aminoacetylated polyvinyl alcohol; the polysulfonedialkylammonium salts; basic proteins, poly-(L)-lysine,poly-(L)-arginine, poly(ornithine), basic gelatins (B-gelatins),chitosan; chitosan with various degrees of acetylation; starch, amylose,amylopectin, cellulose, guar, gum arabic, karaya gum, guar gum, dextran,pullulan, xanthan, curdlan, gellan, carubin, agarose, as well as chitinand chitosan derivatives having the following functional groups invarious degrees of substitution: 2-aminoethyl, 3-aminopropyl,2-dimethylaminoethyl, 2-diethylaminoethyl, 2-diisopropylaminoethyl,2-dibutylaminoethyl, 3-diethylamino-2-hydroxypropyl,N-ethyl-N-methylaminoethyl, N-ethyl-N-methylaminopropyl,2-diethylhexylaminoethyl, 2-hydroxy-2-diethylaminoethyl,2-hydroxy-3-trimethylammonionopropyl,2-hydroxy-3-triethylammonionopropyl, 3-trimethylammonionopropyl,2-hydroxy-3-pyridiniumpropyl and S,S-dialkylthioniumalkyl. Thesederivatives may additionally comprise nonionic functional groups invarious degrees of substitution, such as methyl, ethyl, propyl,isopropyl, 2-hydroxymethyl, 2-hydroxypropyl and 2-hydroxybutyl groups,for example, and also esters with aliphatic carboxylic acids (C₂ toC₁₈); and also n,m-ionenes, poly(aniline); poly(pyrrole);poly(viologens) and also poly(amidoamines) based on piperazine.

II. Amphiphilic Copolymer

A preferred amphiphilic copolymer (nonionic copolymer) component for usein the methods and compositions in the present invention is a copolymerof an ionizable monomer and a hydrophobic monomer. The amphiphiliccopolymer is preferably comprised of a nonionic hydrophilic monomer andnonionic hydrophobic monomer. Amphiphilic copolymers for use inconstructing microgels of the present invention are selected fromamphiphilic diblock copolymers, amphiphilic triblock copolymers,amphiphilic multiblock copolymers, and amphiphilic graft copolymers. Theamphiphilic copolymer is preferably a di- or triblock copolymer. Theamphiphilic copolymer is preferably comprised of (poly(ethylene oxide)and a monomer selected from the group consisting essentially of(poly(propylene oxide), poly(butylene oxide), polystyrene,polyisobutylene, poly(methyl methacrylate), and poly(tert-butylacrylate)).

Amphiphilic copolymers for use in constructing responsive microgels ofthe present invention generally have a molecular weight in the range offrom about 200 to about 1,000,000, preferably from about 500 to about500,000, and more preferably from about 200 to about 200,000. Theamphiphilic copolymers generally have a hydrophilic/lipophilic balancein the range of from about 0.001 to about 100.

A preferred embodiment of the present invention comprises an amphiphiliccopolymer comprised of a diblock, triblock, or multiblock copolymer,preferably a diblock or triblock copolymer, more preferably a diblockcopolymer. A particularly preferred embodiment comprises a triblockcopolymer wherein one block comprises polyoxyethylene. Anotherparticularly preferred embodiment comprises a triblock copolymer whereinone block comprises polyoxypropylene.

Any of the hydrophilic blocks of various chemistry and formula weight ofthe amphiphilic copolymers herein can be used in combination with any ofthe hydrophobic blocks of various chemistry and formula weight tocompose an amphiphilic ‘dangling chain’. The hydrophilic blocks recitedinfra (i.e., A. Hydrophilic Monomers and Polymers) can be used in thecompositions described herein either as an element of the ionizablenetwork (polyelectrolyte) and/or an element of an amphiphilic ‘danglingchain’ copolymer.

The hydrophilic blocks of the amphiphilic diblock, triblock, ormultiblock copolymers can have formula weights in the range from about200 to about 500,000, preferably from about 2,500 to about 250,000, morepreferably from about 500 to about 100,000. The hydrophobic blocks ofthe amphiphilic diblock, triblock, or multiblock copolymers useful inthe present invention can have formula weights in the range of fromabout 1,000 to about 500,000, preferably from about 2,500 to about250,000, more preferably from about 500 to about 100,000.

Amphiphilic graft copolymers useful in the present invention possessrotatable side chain block regions that can rotate or fold to becomepart of the aggregates within the microgels of the present invention.The number of side chains present in each of the amphiphilic graftcopolymers can be in the range of from about 1 to about 10000. Theformula weights of the various blocks in the amphiphilic copolymers canbe varied independently of each other.

A. Hydrophilic Monomers and Polymers

Examples of monomer repeat units that can be used in the preparation ofhydrophilic blocks of the amphiphilic copolymer (or as monomers of theionizable network) are set forth as follows. Poly(acrylic acid) andpoly(metal acrylates) are preferred.

1. Example Monomer Units Useful as Repeat Units in Hydrophilic BlocksPolyacrylic acid Poly(metal acrylate), M = Li, Na, K, Cs PolyacrylamidePoly(methacrylic acid), R = H, alkyl Poly(metal methacrylate)Polymethacrylamide M = Li, Na, K, Cs R = H, alkyl Polystyrene sulfonicacid Polystyrene sulfonic acid metal salt, M = Li, Na, K, Cs Polystyrenecarboxylic Polystyrene carboxylic acid, metal acid salt M = Li, Na, K,Cs Poly(vinyl alcohol), R = H, alkyl Poly(4-vinyl-N- R = H, alkylalkyllpyridinium halide), Poly(2-vinyl-N-alkyllpyridiniumhalide)\Poly(hydroxyethyl methacrylate) Poly(itaconic acid)Poly(N,N,N-trialkyl-4-vinylphenylammonium halide)Poly(N,N,N-trialkyl-4-vinylbenzylammonium halide) Percent quaternization10% to 70% Poly(N,N,N-trialkyl-4-vinylphenethylammonium halide)Poly(L-glutamic acid) Poly(L-aspartic acid) Hyaluronic acid

Amino acids used to compose hydrophilic blocks of the amphiphiliccopolymer: Serine Threonine Tyrosine Lysine Arginine Histidine Asparticacid Glutamic acid

2. Example Polymers Useful as Hydrophilic Blocks

Polymers as hydrophilic blocks of the nonionic copolymer (amphiphiliccopolymer) for employment in the ‘dangling chains’ of the responsivemicrogel of the present invention also include, but are not limited to:

Poly(sodium 1-carboxylatoethylene), Poly(5-hydroxy-1-pentene),5,8-poly-5,7-dodecadiynediol, 10,13-poly-10,12-heptacosadiynoic acid,2,5-poly-2,4-hexadienedioic acid, 2,5-poly-2,4-hexadienoic acid,(6-amino)-2,5-poly-2,4-hexadienoic acid,(6-amino)2,5-poly-2,4-hexadienoic acid, hydrochloride,2,5-poly-2,4-hexadiynediol, 10,13-poly-10,12-nonacosadiynoic acid,2,5-poly-2,4,6-octatriynediol, 10,13-poly-10,12-pentacosadiynoic acid,2,5-poly-5-phenyl-2,4-pentadienoic acid, Poly(2-aminoisobutyric acid),dichloroacetic acid complex, Poly(L-arginine), Poly(L-nitroarginine),Poly(L-aspartic acid), Poly(beta-benzyl-L-aspartic acid),Poly[beta-(p-chloro-benzyl)-L-aspartic acid], Poly(beta-ethyl-L-asparticacid), Poly[beta-(2-phenyl-ethyl)-L-aspartic acid],Poly(alpha-isobutyl-L-aspartic acid),Poly(beta-N-propyl-L-asparticacid), Poly(2,4-diaminobutyricacid),Poly(N-benzyloxycarbonyl-2,4-diaminobutyric acid), Poly(D-glutamicacid), Poly(gamma-benzyl-D-glutamic acid),Poly(gamma-m-chloro-benzyl-D-glutamic acid),Poly(gamma-o-chloro-benzyl-D-glutamic acid),Poly(gamma-p-chloro-benzyl-D-glutamic acid),Poly(gamma-methyl-D-glutamic acid),Poly(gamma-phthalimidomethyl-L-glutamic acid),Poly(gamma-N-amyl-L-glutamic acid), Poly(gamma-benzyl-L-glutamic acid),Poly(gamma-m-chloro-benzyl-L-glutamic acid),Poly(gamma-o-chloro-benzyl-L-glutamic acid),Poly(gamma-p-chloro-benzyl-L-glutamic acid),Poly(gamma-N-butyl-L-glutamic acid), Poly(gamma-N-dodecyl-L-glutamicacid), Poly(gamma-N-ethyl-L-glutamic acid),Poly[gamma-N-(2-chloro-ethyl)-L-glutamic acid],Poly[gamma-N-(2-phenyl-ethyl)-L-glutamic acid],Poly(gamma-N-hexyl-L-glutamic acid), Poly(gamma-methyl-L-glutamic acid),Poly(gamma-methyl-L-glutamic acid), dimethyl phthalate complex,Poly(gamma-N-octyl-L-glutamicacid), Poly(gamma-N-propyl-L-glutamicacid), Poly[gamma-N-(3-phenyl-propyl)-L-glutamic acid],Poly(L-glutamine), Poly[N5-(4-hydroxybutyl)-L-glutamine],Poly[N5-(2-hydroxyethyl)-L-glutamine],Poly[N5-(3-hydroxypropyl)-L-glutamine], Poly(D-glutamyl-L-glutamicacid), Poly(gamma-benzyl-D-glutamyl-L-glutamic acid),Poly(gamma-ethyl-D-glutamyl-L-glutamic acid),Poly[gamma-(2-phenyl-ethyl)-D-glutamyl-L-glutamic acid],Poly(L-histidine), Poly(1-benzyl-L-histidine), Poly(L-histidine),hydrochloride, Poly(gamma-hydroxy-L-alpha-aminoveleric acid),Poly(L-lysine), Poly(E-benzyloxycarbonyl-L-lysine), Poly(L-lysine),hydrobromide, Poly(L-methionine-s-carboxymethylthetin),Poly(L-methionine-s-methylsulfonium bromide), Poly(L-serine),Poly(gamma-hydroxy-L-proline), Poly(hydroxymethylene),Poly(1-hydroxytrimethylene),Poly(3,3-bishydroxymethyltrimethyleneoxide),Poly(3-hydroxytrimethyleneoxide), Poly(vinyl alcohol), Poly(ethyleneglycol), Poly(2-methyl-vinyl alcohol), Poly(hydroxymethylene),Poly(cinnamic acid), Poly(crotonic acid), Poly(3-bromo acrylic acid),Poly(3-ethyl acrylic acid), Poly(N-acetyl-alpha-amino acrylic acid),Poly(alpha-bromoacrylic acid), Poly(alpha-chloroacrylic acid),Poly(alpha-fluoroacrylic acid), Poly(sodium alpha-chloroacrylate),Poly(3-oxa-5-hydroxypentyl methacrylate), Poly(2-hydroxyethyl acrylate),Poly(2-hydroxypropyl acrylate), Poly(beta-chloro-2-hydroxypropylacrylate), Poly[N-(2-hydroxyethyl)-3,6-dichlorocarbazolyl acrylate],Poly[N-(2-hydroxyethyl)carbazolyl acrylate],Poly(acryloyl-beta-hydroxyethyl-3,5-dimitrobenzoat),Poly(methacryloyl-beta-hydroxyethyl-3,5-dimitrobenzoat),Poly(N-(2-hydroxyethyl)carbazolyl methacrylate), Poly(2-hydroxyethylmethacrylate), Poly(2-hydroxypropyl methacrylate),Poly(3-methoxy-2-hydroxypropyl methacrylate),Poly[1-(2-hydroxyethyl)pyridiniumbenzene sulfonate methacrylate],Poly[1-(2-hydroxyethyl)trimethylamoniumbenzene sulfonate methacrylate],Poly[N-(2-hydroxyethyl)phthalimido methacrylate],Poly[N-(hydroxyethyl)carbazolyl methacrylate],Poly(N-ethyl-3-hydroxymethylcarbazolyl methacrylate), Poly(2-sulfonicacid-ethyl methacrylate), Poly(2-trimethylammonium ethyl methacrylatechloride), Poly(2-trimethylammoniummethyl methacrylate chloride),Poly(methacrylonitrile), Poly(thiolacrylic acid), Poly(acrylonitrile),Poly(acrylamide), Poly(methacrylamide), Poly(N,N-dimethylacrylamide),Poly[(N-methylol)acrylamide], Poly[N-methoxymethyl methacrylamide],Poly(N-methyl methacrylamide), Poly(N-2-methoxyethyl methacrylamide),Poly[N-(2-hydroxypropyl)methacrylamide], Poly(2-methylpropanesulfonatesodium 2-acrylamido), Poly(2-methylpropanesulfonic acid 2-acrylamido),Poly[(p-amino)-styrene], Poly[4-(4-hydroxybutoxymethyl)styrene],Poly[4-(2-hydroxyethoxymethyl)styrene],Poly[4-(2-hydroxyiminoethyl)styrene],Poly[4-(1-hydroxyiminoethyl)styrene], Poly[4-(n-2-hydroxybutyl)styrene],Poly[4-(1-hydroxy-3-dimethylaminopropyl)styrene],Poly[4-(1-hydroxy-1-methylbutyl)styrene],Poly[4-(1-hydroxy-1-methylethyl)styrene],Poly[4-(1-hydroxy-1-methylhexyl)styrene],Poly[4-(1-hydroxy-1-methylpentyl)styrene],Poly[4-(1-hydroxy-1-methylpropyl)styrene], Poly(2-hydroxymethylstyrene),Poly(3-hydroxymethylstyrene), Poly(4-hydroxymethylstyrene),Poly(4-hydroxystyrene), Poly[p-1-(2-hydroxybutyl)-styrene],Poly[p-1-(2-hydroxypropyl)-styrene],Poly[p-2-(2-hydroxypropyl)-styrene],Poly[4-(1-hydroxy-3-morpholinopropyl)styrene],Poly[4-(1-hydroxy-3-piperidinopropyl)styrene],Poly(p-octylaminesulfonate styrene), Poly(2-carboxystyrene),Poly(4-carboxystyrene), Poly(styrene sulfonic acid), Poly(vinyl sulfonicacid), Poly[N-(2-hydroxypropyl)methacrylamide],Poly[oxy(hydroxyphosphinylidene)], Poly(9-vinyladenine), Poly(vinylcarbanilate), Poly(vinylpyrrolidone), Poly(vinyl succinamic acid),Poly(N-isopropylacrylamide), Poly(methacrylic acid), Poly(itaconicacid), Poly(glycidyl methyl itaconate), Poly(monomethyl itaconate),Poly[N-(p-chlorophenyl)itaconimide], Poly[N-(p-tolyl)itaconimide],Poly[N-(2-chloroethyl)itaconimide],Poly[N-(4-acetoxyphenyl)itaconimide],Poly[N-(4-chlorophenyl)itaconimide],Poly[N-(4-ethoxycarbonylphenyl)itaconimide], Poly(N-benzylitaconimide),Poly(N-butylitaconimide), Poly(N-ethylitaconimide),Poly(N-isopropylitaconimide), Poly(N-isobutylitaconimide),Poly(N-methylitaconimide), Poly(N-naphthylitaconimide),Poly(N-phenylitaconimide), Poly(N-propylitaconimide),Poly(N-tolylitaconimide), Poly(alpha-chlorovinyl acetic acid),Poly(carboxychloromethyl ethylene), Poly(4-vinyl phenol),Poly(o-hydroxy-vinylphenylketone), Poly(alpha-phenylvinyl phosphonicacid),Poly[(1,2,5-trimethyl-4,4i-hydroxypyridiumchlorideethynyl)ethylene],Poly(allyl alcohol), Poly(acrylic acid), Poly[2-(3-sodiumsulfonato-2-methylpropyl)methacrylamide], Poly(3-sodium sulfonatopropylmethacrylate), Poly(3-oxa-5-hydroxypentyl methacrylate),Poly(diethylenegycol dimethacrylate), Poly(trimethyleneglycoldimethacrylate), Poly(triethyleneglycol dimethacrylate),Poly(ethyleneglycol N-phenylcarbamate methacrylate),Poly(acryloyl-L-glutamic acid), Poly(methacryloyl-L-glutamic acid),Poly(butadiene-1-carboxylic acid), Poly(crotonate acid),Poly(trans-4-ethoxy-2,4-pentadienoic acid), Poly(alpha-phenylvinylphosphonic acid), Poly(vinylbenzoic acid), Poly(2-acryloyloxy benzoicacid), Poly[1-(2-hydroxyethylthio)-1,3-butadiene], Poly(2,5-dicarboxylicacid-1-hexene), Poly(3-hydroxyisoprene), Poly(alpha-phenylvinylphosphonic acid), Poly(2-chloro-3-hydroxy propene),Poly(2-p-vinylphenylpropanol), Poly(o-hydroxy-vinylphenylketone),Poly(1-vinyl-3-benzyl-imidazolium chloride),Poly(4-vinylbenzyltrimethylammonium chloride),Poly(4-vinylbenzyldimethyl vinylbenzyl ammonium chloride),Poly(4-vinylbenzyldimethyl methacryloyl ammonium chloride),Poly(4-vinylbenzyldimethyl acryloyl ammonium chloride),Poly(4-vinylbenzyldimethyl allyl ammonium chloride),Poly(4-vinylphenyltrimethylammonium chloride), Poly(4-vinylphenyldimethyl vinylbenzyl ammonium chloride), Poly(4-vinylphenyl dimethylmethacryloyl ammonium chloride), Poly(4-vinylphenyl dimethyl acryloylammonium chloride), Poly(4-vinylphenyl dimethyl allyl ammoniumchloride), Poly(4-vinylphenethyltrimethylammonium chloride),Poly(4-vinylphenethyldimethyl vinylbenzyl ammonium chloride),Poly(4-vinylphenethyldimethyl methacryloyl ammonium chloride),Poly(4-vinylphenethyldimethyl acryloyl ammonium chloride),Poly(4-vinylphenethyldimethyl allyl ammonium chloride), Poly(vinylacetate), Poly(vinyl butyral), Poly(acetaldehyde), Poly(ethylene oxide),Poly(2-cyanoethyloxymethylene oxide), Poly[(methoxymethyl)ethyleneoxide], Poly(methylene sulfide), Poly(ethylene disulfide), Poly(ethylenesulfide), Poly(ethylene tetrasulfide), Poly(methylene disulfide),Poly(trimethylene disulfide), Poly(ethylene amine), Poly(propyleneamine), Poly(4-vinyl-N-methylpyridinium chloride),Poly(4-vinyl-N-ethylpyridinium chloride),Poly[4-(2-dimethylaminoethoxycarbonyl)styrene], hydrochloride,Poly(4-vinylpyridine), hydrogen chloride,Poly(4-vinyl-N-vinylbenzylpyridinium chloride),Poly(4-vinyl-N-methacryloylpyridinium chloride),Poly(4-vinyl-N-acryloylpyridinium chloride),Poly(4-vinyl-N-allylpyridinium chloride),Poly(2-vinyl-N-methylpyridinium chloride),Poly(2-vinyl-N-ethylpyridinium chloride),Poly(2-vinyl-N-vinylbenzylpyridinium chloride),Poly(2-vinyl-N-methacryloylpyridinium chloride),Poly(2-vinyl-N-acryloylpyridinium chloride),Poly(2-vinyl-N-allylpyridinium chloride), and Poly(2-vinylpyridine)hydrogen chloride.

B. Hydrophobic Monomers and Polymers

The hydrophobic blocks of the amphiphilic diblock, triblock, ormultiblock copolymers useful in the present invention can have formulaweights in the range of from about 500 to about 500,000, preferably fromabout 500 to about 250,000, more preferably from about 500 to about100,000. Examples of monomer repeat units that can be used in thepreparation of hydrophobic blocks are set forth as follows.

1. Example Monomers Units Useful as Repeat Units in Hydrophobic Blocksor Hydrophobic Repeat Units Poly(2-vinylnaphthalene) Poly(caprolactam),R = H, CH₃, alkyl, or aryl group Polystyrene Poly(amide)poly(p-X-styrene), X = alkyl, CH₃, t-Bu, O CH₃, CH₂ Cl, Cl, CN, CHOpoly(alpha-methylstyrene) poly(4-vinylpyridine) poly(2-vinylpyridine)polybutadiene polybutadiene 1,4-addition 1,2-addition polyisoprenepolychloroprene polyethylene polypropylene polyvinylchloridepolyvinylidenechloride polyvinylfluoride polyvinylidenefluoridepolyhexafluoropropene polypropyleneoxide polypropyleneoxidepoly(N-vinylcarbazol) polytetrafluoroethane polysiloxane polyacrylates R= CH₃, alkyl or aryl group R′ = CH₃, any alkyl or aryl group R′ = CH₃,alkyl or aryl group R = CH₃, CH₂ CH₃, t-Butyl, any alkyl or aryl group

Amino acids used to compose hydrophobic blocks of the amphiphiliccopolymer: Alanine Valine Leucine Tryptophan Phenylalanine MethionineProline

2. Example Polymers Useful as Hydrophobic Blocks

Polymers as hydrophobic blocks of the nonionic copolymer (amphiphiliccopolymer) for employment in the ‘dangling chains’ of the responsivemicrogel of the present invention also include, but are not limited to:

Poly[thio(2-chlorotrimethylene)thiotrimethylene],Poly[thio(1-iodiethylene)thio(5-bromo-3-chloropentamethylene),Poly[imino(1-oxoethylene)silylenetrimethylene],Poly(oxyiminomethylenehydrazomethylene),Poly[oxy(1,1-dichloroethylene)imino(1-oxoethylene)],Poly[(6-chloro-1-cyclohexen-1,3-ylene)-1-bromoethylene],Poly[(dimethylimino)ethylenebromide],Poly[(oxycarbonyloxymethyl)ethylene], Poly(1,1-dimethylethylene),Poly(1-methyl-1-butenylene),Poly[(2-propyl-1,3-dioxane-4,6-diyl)methylene],Poly[1-(methoxycarbonyl)ethylene], Poly(glycyl-6-aminocarproic acid),Poly(glycyl-6-aminocarproic acid-3-amino-propionic acid),Poly(L-alanyl-4-aminobutyric acid), Poly(L-alanyl-6-aminocaproic acid),Poly(L-alanyl-3-aminopropionic acid), Poly(L-alanyl-5-aminovalericacid), Poly(2-aminocyclopentylenecarboxy acid),Poly(2-aminoethylenesulfonic acid), Poly(3-aminopropionic acid),Poly(1-methyl-3-aminopropionic acid),Poly[(3-aminocyclobutylene)-propionic acid],Poly[(2,2-dimethyl-3-aminocyclobutylene)-propionic acid],Poly(2-aminoisobutyric acid), Poly(3-aminobutyric acid),Poly(4-aminobutyric acid), Poly(5-aminovaleric acid),Poly(6-aminocaproic acid), Poly(D-(−)-3-methyl-6-aminocaproic acid),Poly(6-methyl-6-aminocaproic acid), Poly(6-aminothiocaproic acid),Poly(7-aminoenanthic acid), Poly((R)-3-methyl-7-aminoenanthic acid),Poly((S)-4-methyl-7-aminoenanthic acid),Poly((R)-5-methyl-7-aminoenanthic acid),Poly((R)-6-methyl-7-aminoenanthic acid), Poly(N-methyl-7-aminoenanthicacid), Poly(7-aminothioenanthic acid), Poly(8-aminocaprylic acid),Poly(9-aminopelargonic acid), Poly(10-aminocapric acid),Poly(11-aminoundecanoic acid), Poly(N-allyl-11-aminoundecanoic acid),Poly(N-ethyl-11-aminoundecanoic acid), Poly(2-methyl-11-aminoundecanoicacid), Poly(N-methyl-11-aminoundecanoic acid),Poly(N-phenyl-11-aminoundecanoic acid),Poly(N-piperazinyl-11-aminoundecanoic acid), Poly(12-aminolauricacid),Poly(aminoformicacid), Poly(N-butyl-aminoformic acid),Poly(2-methyl-N-butyl-aminoformic acid), Poly(N-phenyl-aminoformicacid), Poly[imino-(1-oxo-2,3-dimethyltrimethylene)],Poly[imino-(1-oxo-3-ethyltrimethylene)],Poly[imino-(1-oxo-4-methylhexamethylene)],Poly[imino-(1-oxo-3-methylhexamethylene)],Poly[imino-(1-oxo-5-methylhexamethylene)],Poly[imino-(1-oxo-3-methyl-6-isopropylhexamethylene)],Poly[imino-(1-oxo-3-methyltrimethylene)],Poly[imino-(1-oxo-3-vinyltrimethylene)],Poly[N-(2-methylbutyl)iminocarbonyl],Poly[N-(phenylpropyl)iminocarbonyl], Poly(N-methyldodecanelactam),Poly(L-alanine), Poly(beta-L-alanine), Poly(N-methyl-L-alanine),Poly(L-phenylalanine), Poly(2-butyl-2-methyl-beta-alanine),Poly(2,2-dimethyl-beta-alanine), Poly(3,3-dimethyl-beta-alanine),Poly(2-ethyl-2-methyl-beta-alanine),Poly(2-methyl-2-propyl-beta-alanine), Poly(N-isopropyl-beta-alanine),Poly(3-methyl-beta-alanine), Poly(N-methyl-beta-alanine),Poly(N-phenyl-beta-alanine), Poly(mathacryloyl-D-alanine),Poly(M-methacryloyl-L-alanine), Poly(L-cysteine), Poly(L-glycine),Poly(L-leucine), Poly(isoleucine), Poly(N-trifluoroacetal-L-lysine),Poly(N-carbobenzoxy-L-lysine), Poly(methionine), Poly(L-tyrosine),Poly(o-acetal-hydroxyproline), Poly(o-acetal-L-serine),Poly(alpha-amino-n-butyric acid), Poly(s-carbobenzoxymethyl-L-cysteine),Poly(3,4-dihydro-L-proline), Poly(o-p-tolylsulfonyloxy-L-proline),Poly(gamma-hydroxy-o-acetyl-L-alpha-aminoveleric acid), Poly(L-valine),Poly(L-proline), Poly(L-proline), acid complex, Poly(L-proline), aceticacid complex, Poly(L-proline), formic acid complex, Poly(L-proline),propionic acid complex, Poly(o-acetyl-hydroxy-L-proline),Poly(o-acetyl-L-serine), Poly(o-benzyloxycarbonyl-L-tyrosine),Poly(s-benzyloxycarbonyl-L-cysteine), Poly(s-benzylthio-L-cysteine),Poly(methylphosphinidene-trimethylene), Polymalonate, Polysuccinate,Polyglutarate, Polyadipate, Poly(methylene), Poly(diphenylmethylene),Poly(di-p-tolyl-methylene), Poly(ethylene),Poly(chlorotrifluoroethylene), Poly(1-butoxy-2-methyl-ethylene),Poly(1-t-butoxy-2-methyl-ethylene), Poly(1-ethoxy-2-methoxy-ethylene),Poly(1-ethoxy-2-methyl-ethylene), Poly(1-isobutoxy-2-methyl-ethylene),Poly(1-isopropoxy-2-methyl-ethylene), Poly(1-methoxy-2-methyl-ethylene),Poly(1-methyl-2-propoxy-ethylene), Poly(tetrafluoroethylene),Poly(trifluoroethylene), Poly(butylethylene), Poly(t-butylethylene),Poly(cyclohexylethylene), Poly(2-cyclohexylethylene),Poly[(cyclohexylmethyl)ethylene], Poly(3-cyclohexylpropylethylene),Poly(decylethylene), Poly(dodecylethylene), Poly(isobutyl ethylene),Poly(neopentylethylene), Poly(4,4-dimethylpentylethylene),Poly(nonylethylene), Poly(octylethylene), Poly(propylethylene),Poly(propyl-2-propylene), Poly(tetradecylethylene), Poly(vinylbromide),Poly(N-vinyl carbazole), Poly(vinyl chloride), Poly(vinyl fluoride),Poly(vinylidene bromide), Poly(vinylidene chloride),Poly(vinylidenefluoride), Poly(vinylcyclobutane),Poly(vinylcycloheptane), Poly(vinylcyclohexane),Poly(o-methoxy-vinylcyclohexane), Poly(3-methyl-vinylcyclohexane),Poly(4-methyl-vinylcyclohexane), Poly(vinylcyclohexene),Poly(vinylcyclohexylketone), Poly(vinylcyclopentane),Poly[3-(2-vinyl)-6-methyl pyridazinone],Poly[3-(2-vinyl)-6-methyl-4,5-pyridazinone],Poly(cyclopentylmethylethylene), Poly(heptylethylene),Poly(hexyldecylethylene), Poly(hexylethylene), Poly(cyclohexylethylene),Poly(cyclopentylethylene), Poly(cyclopropylethylene),Poly(isopentylethylene), Poly(isopropylethylene),Poly(3,3-dimethylbutylethylene), Poly(isohexylethylene),Poly(1,1-dimethylethylene), Poly(benzylethylene),Poly(N-carbazoylylethylene), Poly(ferrocenylethylene),Poly(indazol-2-ylethylene),Poly[dimethylamino(ethoxy)phosphinylethylene],Poly[dimethylamino(phenoxy)phosphinylethylene],Poly(4,4-dimethyl-oxazolonylethylene),Poly(4,4-dimethyl-oxazolonyl-2-propylene),Poly[(2-methyl-5-pyridyl)ethylene], Poly[(2-methyl-6-pyridyl)ethylene],Poly(2,4-dimethyl-1,3,5-triazinylethylene), Poly(1-naphthylethylene),Poly(2-naphthylethylene), Poly(phenethylethylene),Poly(phenethylmethylethylene), Poly(phenylacetylene),Poly(diphenylphosphinylethylene), Poly(phenylvinylene),Poly(phthalimidoethylene), Poly(2-pyridylethylene),Poly(4-pyridylethylene), Poly(N-pyrrolidinylethylene),Poly(m-tolylmethylethylene), Poly(o-tolylmethylethylene),Poly(p-tolylmethylethylene), Poly(vinyltrimethylgerm anium),Poly(vinylcyclopropane), Poly(N-vinyldiphenylamine),Poly(1-vinylene-3-cyclopentylene), Poly(o-hydroxy-vinylphenylketone),Poly(3-vinyl pyrene), Poly(2-vinylpyridine), Poly(4-vinylpyridine),Poly(2-vinyl-5-methylpyridine), Poly(2-vinyl-5-ethylpyridine),Poly(1-cyano-2-phenylvlnylene), Poly(vinyl 3-trimethylsilylbenzoat),Poly(vinylfuran), Poly(vinylindole), Poly(2-vinyltetrahydrofuran),Poly(N-vinylphthalimide), Poly(1-vinylimidazlo), Poly(1-vinyl-2-methylimidazole), Poly(5-vinyl-2-methylpyridine), Poly(1-vinylnaphthalene),Poly(2-vinylnaphthalene), Poly(5-vinyl-2-picoline), Poly(3-vinylpyrene),Poly(2-vinylpyridine), Poly(4-vinylpyridine),Poly(2-methyl-5-vinylpyridine), Poly(N-vinylcarbazole),Poly(1-vinylnaphthalene), Poly(styryl pyridine), Poly(N-vinylsuccinimide), Poly(1,3-divinyl-imidazolid-2-one),Poly(1-ethyl-3-vinyl-imidazolid-2-one), Poly(p-vinyl benzophenone),Poly(vinyl N,N-diethyl-carbamate), Poly(vinyl cymantrene),Poly[vinyl-tris(trimethoxysiloxy)silane], Poly(alpha-chlorovinyltriethoxysilane), Poly(p-vinylbenzylethylcarbinol),Poly(p-vinylbenzylmethylcarbinol), Poly(divinylaniline),Poly(vinylferrocene), Poly(9-vinylanthracene),Poly(vinylmercaptobenzimidazole), Poly(vinylmercaptobenzoxazole),Poly(vinylmercaptobenzothiazole), Poly(p-vinyl benzophenone),Poly(2-vinyl quinoline), Poly(vinylidene cyanide),Poly(1,2,5-trimethyl-vinylethylnyl-4-piperidinol),Poly(2-vinyl-1,1-dichlorocyclopropane),Poly(2-vinyl-2-methyl-4,4,6,6-tetraphenylcyclotrisiloxane),Poly(N-vinyl-N-methylacetamide), Poly(triethoxysilyl ethylene),Poly(trimethoxysilyl ethylene), Poly(1-acetoxy-1-cyanoethylene),Poly(1,1-dichloroethylene), Poly(1,1-dichloro-2-fluoroethylene),Poly(1,1-dichloro-2,2-difluoroethylene),Poly(1,2-dichloro-1,2-difluoroethylene),Poly[(pentafluoroethyl)ethylene], Poly(tetradecafluoropentylethylene),Poly(hexafluoropropylene), Poly(2,3,3,3-tetrafluoropropylene),Poly(3,3,3-trifluoropropylene), Poly[(heptafluoropropyl)ethylene],Poly(2-iodoethylethylene), Poly(9-iodononylethylene),Poly(3-iodopropylethylene), Poly[(2-acetoxybenzoyloxy)ethylene],Poly(4-acetoxybenzoyloxyethylene),Poly[(1-acetylindazol-3-ylcarbonyloxy)ethylene],Poly(4-benzoylbutyryloxyethylene), Poly(3-bromobenzoyloxyethylene),Poly(4-bromobenzoyloxyethylene), Poly[(t-butoxycarbonylamino)ethylene],Poly(4-t-butylbenzoyloxyethylene), Poly(4-butyryloxybenzoyloxyethylene),Poly(2-chlorobenzoyloxyethylene), Poly(3-chlorobenzoyloxyethylene),Poly(4-chlorobenzoyloxyethylene), Poly(cyclohexanoyloxyethylene),Poly(cyclohexylacetoxyethylene), Poly(4-cyclohexylbutyryloxyethylene),Poly(cyclopentanoyloxyethylene), Poly(cyclopentyl acetoxyethylene),Poly(4-ethoxybenzoyloxyethylene), Poly(4-ethylbenzoyloxyethylene),Poly[(2-ethyl-2,3,3-trimethylbutyryloxy)ethylene],Poly(trifluoroacetoxyethylene), Poly(heptafluorobutylryloxyethylene),Poly[(undecafluorodecanoyloxy)ethylene],Poly[(nonadecafluorodecanoyloxy)ethylene],Poly[(undecafluorohexanoyloxy)ethylene],Poly[(pentadecafluorooctanyloxy)ethylene],Poly[(pentafluoropropionyloxy)ethylene],Poly[(heptafluoroisopropoxy)ethylene], Poly(formyloxyethylene),Poly(isonicotinoyloxyethylene), Poly(4-isopropylbenzoyloxyethylene),Poly[(2-isopropyl-2,3-dimethylbutyryloxy)ethylene],Poly[(2-methoxybenzoyloxy)ethylene],Poly[(3-methoxybenzoyloxy)ethylene],Poly[(4-methoxybenzoyloxy)ethylene], Poly[(2-methylbenzoyloxy)ethylene],Poly[(3-methylbenzoyloxy)ethylene], Poly[(4-methylbenzoyloxy)ethylene],Poly[(1-methylcyclohexanoyloxy)ethylene],Poly(3,3-dimethyl-3-phenylpropionyloxyethylnene),Poly[(3-trimethylsilylbenzoyloxy)ethylene],Poly[(4-trimethylsilylbenzoyloxy)ethylene),Poly[(2,2-dimethylvaleryloxy)ethylene],Poly[(2,2,3,3-tetramethylvaleryloxy)ethylene],Poly[(2,2,3,4-tetramethylvaleryloxy)ethylene],Poly[(2,2,4,4-tetramethylvaleryloxy)ethylene],Poly(nicotinoyloxyethylene), Poly(nitratoethylene),Poly[(3-nitrobenzoyloxy)ethylene], Poly[(4-nitrobenzoyloxy)ethylene],Poly[(4-phenylbenzoyloxy)ethylene], Poly(pivaloyloxyethylene),Poly[(4-propionyloxybenzoyloxy)ethylene], Poly(propionyloxyethylene),Poly[(4-p-toluoylbutyryloxy)ethylene],Poly[(1,2-diethoxycarbonyl)ethylene],Poly[(1,2-dimethoxycarbonyl)ethylene],Poly[(1,2-dipropoxycarbonyl)ethylene],Poly(2-bromotetrafluoroethyliminotetrafluoroethylene),Poly[(biphenyl-4-yl)-ethylene], Poly(2-chloroethoxyethylene),Poly(hexadecyloxyethylene), Poly(isobutoxyethylene),Poly(1-methoxycarbonyl-1-phenylethylene), Poly(9-acrydinylethylene),Poly(4-methoxybenzylethylene), Poly[(3,6-dibromocarbazoyl)ethylene]Poly(propylene oxide) Poly(dimethylpentylsilylethylene),Poly(3,5-dimethylpyrozoylylethylene),Poly(2-diferrocenyl-furyl-methylene), Poly(ethoxyoxaloyloxymethylethylene), Poly(9-ethyl-3-carbazoyl ethylene), Poly(fluorenylethylene),Poly(imidazoethylene), Poly[(8-methoxycarbonyloctyl)ethylene],Poly(1-methoxy-4-naphthyl ethylene), Poly(2-methyl-5-pyridyl ethylene),Poly(propoxyoxaloyloxymethyl ethylene),Poly(1,1-diphenyl-2-vinylcyclopropane), Poly(p-anthrylphenylethylene),Poly[1-(N-ethyl-N-(1,4,7,10,13-pentaoxacyclopentadecyl)-carbamoyl)ethylene],Poly(N-carbazolylcarbonyl ethylene), Poly(morpholinocarbonyl ethylene),Poly(piperidinocarbonyl ethylene), Poly(N-benztriazolylethylene),Poly[6-(N-carbazoyl)hexyl ethylene],Poly(2,4-dimethyl-6-triazinylethylene),Poly(diphenylthiophosphinylideneethylene),Poly(2-methyl-5-pyridylethylene), Poly(N-thiopyrrolidonylethylene),Poly(N-1,2,4-triazolylethylene), Poly(phenothiazinyl ethylene),Poly(L-menthyloxycarbonylaminoethylene), Poly(N-3-methyl-2-pyrrolidoneethylene), Poly(p-vinyl-1,1-diphenyl ethylene),Poly(S-vinyl-O-ethylthioacetal formaldehyde), Poly(N-vinylphthalimide),Poly[N-(4-vinylphenyl)phthalimide],Poly[2-methyl-5-(4′-vinyl)phenyltetrazole],Poly[5-phenyl-2-(4′-vinyl)phenyltetrazole],Poly(N,N-methyl-vinyltoluenesulfonamide), Polyallene, Poly(1-butene),Poly(1-bromo-1-butene), Poly(1-butyl-1-butene),Poly(1-t-butyl-1-butene), Poly(1-chloro-1-butene),Poly(2-chloro-1,4,4-trifluoro-1-butene), Poly(1-decyl-1-butene),Poly(1-ethyl-butene), Poly(1,4,4-trifluoro-1-butene),Poly(octafluoro-1-butene), Poly(1-heptyl-1-butene),Poly(4-p-chlorophenyl-1-butene), Poly(4-p-methoxyphenyl-1-butene),Poly(4-cyclohexyl-1-butene), Poly(4-phenyl-1-butene), Poly(2-butene),Poly(isoprene), Poly(3-acetoxyisoprene), Poly(1-isopropyl-1-butene),Poly[3-(1-cyclohexenyl)isopropenyl acetate], Poly(4-methoxy-1-butene),Poly(4-methoxycarbonyl-3-methyl-1-butene), Poly(1,2-dimethyl-butene),Poly(1-phenyl-butene), Poly(1-propyl-butene),Poly[(3-methyl)-1-butene)], Poly[(4-methyl)-1-butene)],Poly[(4-phenyl)-1-butene)], Poly[(4-cyclohexyl)-1-butene)],Poly[(4-N,N-diisopropylamino)-1-butene)],Poly[(3,3-dimethyl)-1-butene)], Poly[(3-phenyl)-1-butene)],Poly[(4-o-tolyl)-1-butene)], Poly[(4-p-tolyl)-1-butene)],Poly[(4,4,4-trifluoro)-1-butene)], Poly[(3-trifluoromethyl)-1-butene)],Poly[(4-trimethylsilyl)-1-butene], Poly(1,3,3-trimethylbutene),Poly(1,4-p-methoxyphenylbutene), Poly(1,4-p-chlorophenyl butene),Poly(1,4-cycl ohexylbutene), Poly(1,4-phenylbutene), Poly(1,2-diethylbutene), Poly(2,2-dimethylbutene), Poly(1,3-cyclobutylene),Poly[(1-cyano)-1,3-cyclobutylene], Poly(N-butenyl carbazole),Poly(1-decene), Poly(1-docosene), Poly(dodecamethylene),Poly(1,2-chloro-dodecamethylene), Poly(1-methyl-dodecamethylene),Poly(1-dodecene), Poly(1-nonene), Poly(1-heptene),Poly(6,6-dimethyl-1-heptene), Poly(5-methyl-1-heptene),Poly(heptamethylene), Poly(1,2-dichloro-heptamethylene),Poly[(5-methyl)-1-heptene], Poly(1-hexadecene), Poly(1-hexene),Poly[(3-methyl)-1-hexene], Poly[(4-methyl)-1-hexene],Poly[(4,4-dimethyl)-1-hexene], Poly[(4-ethyl)-1-hexene],Poly[(5-methyl)-1-hexene], Poly(1,2-cyclohexylene),Poly(1,2-cyclopentylene-alt-ethylene),Poly(1,3-cyclopentylene-alt-methylene), Poly(isobutene),Poly(1-octadecene), Poly(octamethylene), Poly[(1-methyl)octamethylene],Poly(1-octene), Poly(6,6-dimethyl-4,8-dioxaspiro-1-octene),Poly(1-octadecene), Poly(1-pentene), Poly(cyclopentene),Poly(1,3-dione-4-cyclopentene), Poly(3,3-dimethoxycyclopentene),Poly(1-pentadecene), Poly(5-amino-1-pentene),Poly(5-cyclohexyl-1-pentene), Poly[5-(N,N-dimethyl)amino-1-pentene],Poly[5-(N,N-diisobutyl)amino-1-pentene],Poly[5-(N,N-dipropyl)amino-1-pentene], Poly(4,4-dimethyl-1-pentene),Poly(3-methyl-1-pentene), Poly(3-ethyl-1-pentene),Poly(4-methyl-1-pentene), Poly(5,5,5-trifluoro-1-pentene),Poly(4-trifluoromethyl-1-pentene), Poly(5-trimethylsilyl-1-pentene),Poly(2-methyl-1-pentene), Poly(5-phenyl-1-pentene),Poly(1,2-cyclopentylene), Poly(3-chloro-1,2-cyclopentylene),Poly(pentamethylene), Poly(1,2-dichloropentamethylene),Poly(hexafluoroisobutylene), Poly(chloroprene), Poly(propene),Poly(3-cyclohexylpropene), Poly(3-cyclopentylpropene),Poly(hexafluoropropene), Poly(3-phenylpropane),Poly[3-(2′,5′-dimethylphenyl)propene],Poly(3-(3′,4′-dimethylphenyl)propene],Poly[3-(3′,5′-dimethylphenyl)propene], Poly(3-silylpropene),Poly(3-p-tolylpropene), Poly(3-m-tolylpropene), Poly(3-o-tolylpropene),Poly(3-trimethylsilylpropene), Poly(3,3,3-trifluoropropene),Poly(3,3,3-trichloropropene), Poly(1-chloropropene),Poly(2-chloropropene), Poly(2,3-dichloropropene),Poly(3-chloro-2-chloromethylpropene), Poly(ethyl-2-propylene),Poly(1-nitropropylene), Poly(2-trimethylsilylpropene),Poly[1-(heptafluoroisopropoxy)methylpropylene],Poly[(1-heptafluoroisopropoxy)propylene], Poly(N-propenyl carbazole),Poly(propylidene), Poly(isopropenyltoluene), Poly(1-tridecene),Poly(1-tetradecene), Poly(vinylcyclobutane), Poly(vinylcycloheptane),Poly(vinylcyclohexane), Poly(vinylcyclopentane),Poly(vilnylcyclopropane), Poly(1-vinylene-3-cyclopentylene),Poly(octamethylene), Poly(1-methyloctamethylene), Poly(decamethylene),Poly(1,2-dichloro-decamethylene), Poly(2,5-pyrazinecyclobutylene),Poly(2,4-diphenyl-2,5-pyrazinecyclobutylene), Poly(1-undecene),Poly[(R)(−)-3,7-dimethyl-1-octene], Poly[(S)(+)-5-methyl-1-heptene],Poly[(S)(+)-4-methyl-1-hexene], Poly[(S)(+)-4-methyl-1-hexyne],Poly[(S)(+)-6-methyl-1-octene], Poly[(S)(+)-3-methyl-1-pentene],Poly[(R)-4-phenyl-1-hexene], Poly(dimethyl 2,5-dicarboxylate-1-hexene),Poly[(S)-5-phenyl-1-heptene], Poly(1-ethyl-1-methyltetramethylene),Poly(1,1-dimethyltetramethylene), Poly(1,1-dimethyltrimethylene),Poly(1,1,2-trimethyltrimethylene), Poly(acryloyl chloride),Poly(allylacrylate), Poly(allyl chloride), Poly(allylbenzene),Poly(diallyl phthalate), Poly(diallylcyanamide), Poly(acryloylpyrriolidone), Poly(allylcyclohexane), Poly(N-allylstearamide),Poly(allyl chloroacetate), Poly(allyl glycidyl phthalate),Poly(allylcyclohexane), Poly(allyltriethoxysilane), Poly(allylurea),Poly(allylbenzene), Poly(acetylene), Poly(beta-iodophenylacetylene),Poly(diacetylene), Poly(phenyl acetylene), Poly(3-methyl-1-pentyne),Poly(4-methyl-1-hexyne), Poly(5-methyl-1-heptyne),Poly(6-methyl-1-octyne), Poly(3,4-dimethyl-1-pentyne),Poly(2,3-dihydrofuran), Poly(N,N-dibutylacrylamide),Poly(N-docosylacrylamide), Poly(N-dodecylacrylamide),Poly(N-formylacrylamide), Poly(N-hexadecylacrylamide),Poly(N-octadecylacrylamide), Poly(N-octylacrylamide), Poly(N-phenylacryl amide), Poly(N-propylacryl amide), Poly(N-tetradecylacrylamide),Poly(N-butylacrylamide), Poly(N-sec-butylacrylamide),Poly(N-t-butylacrylamide), Poly(isodecylacrylamide),Poly(isohexylacrylamide), Poly(isononylacrylamide),Poly(isooctylacrylamide), Poly[N-(1,1-dimethyl-3-oxobutyl)acrylamide],Poly[1-oxy-(2,2,6,6-tetramethyl-4-piperidyl)acrylamide],Poly(N,N-dibutylacrylamide), Poly(N,N-diethylacrylamide),Poly(N,N-diisopropylacrylamide), Poly(N,N-diphenylacrylamide),Poly[N-(1,1-dimethyl-3-oxobutyl)acrylamide],Poly[N-(1-methylbutyl)acrylamide], Poly(N-methyl-N-phenylacrylamide),Poly(N-phenyl-N-1-naphthylacrylamide),Poly(N-phenyl-N-2-naphthylacrylamide), Poly(morpholylacrylamide),Poly(N-octadecylacrylamide), Poly(piperidylacrylamide),Poly(4-butoxycarbonylphenyl methacrylamide),Poly(N-t-butylmethacrylamide), Poly(N-benzyl methacrylamide),Poly(N-phenyl methacrylamide), Poly[N-(p-chlorophenyl)methacylamide],Poly[N-(p-methoxyphenyl)methacrylamide],Poly[N-(p-methylphenyl)methacrylamide],Poly[N-(p-nitrophenyl)methacrylamide],Poly[N-(p-stilbenzyl)methacrylamide],Poly[N-(4′-nitro-p-stibenzyl)methacrylamide],Poly(N-phenylmethacrylamide), Poly(1-deoxy-D-glucitol methacrylamide),Poly(4-carboxyphenylmethacrylamide),Poly(4-ethoxycarbonylphenylmethacrylamide),Poly(4-methoxycarbonylphenylmethacrylamide), Poly(N-allylmethacrylamide), Poly[1-(N-carbethoxyphenyl)methacrylamide],Poly(p-ethoxycarbonyl phenylmethacrylamide), Poly(carbethoxyphenylmethacrylamide), Poly(N-methyl-N-alpha-methylbenzyl-acrylamide),Poly(N-propyl-N-alpha-methylbenzyl-acrylamide),Poly(p-acrylamidomethylamino azobenzene), Poly(allyl acrylate),Poly(biphenyloxyhexamethylene acrylate), Poly(n-butylacrylate),Poly(2-nitrobutylacrylate), Poly(sec-butyl acrylate), Poly(t-butylacrylate), Poly(p-carboxyphenyl acrylate), Poly(glycidyl acrylate),Poly(isobutyl acrylate), Poly(isopropyl acrylate), Poly(cresylacrylate), Poly(decylacrylate),Poly(1,1-dihydroperfluoro-decylacrylate), Poly(docosylacrylate),Poly(dodecylacrylate), Poly(hexadecylacrylate), Poly(heptylacrylate),Poly(octadecylacrylate), Poly(octylacrylate),Poly(1,1-dihydroperfluorooctylacrylate), Poly(tetradecylacrylate),Poly(isopropyl acrylate), Poly(benzyl acrylate), Poly(4-biphenylylacrylate), Poly(L-bornyl acrylate), Poly(4-butoxycarbonylphenylacrylate), Poly(2-t-butylphenyl acrylate), Poly(4-t-butylphenylacrylate), Poly[(1-chlorodifluoromethyl)tetrafluoroethyl acrylate],Poly[3-chloro-2,2-bis(chloromethyl)propyl acrylate], Poly(2-chlorophenylacrylate), Poly(4-chlorophenyl acrylate), Poly(2,4-dichlorophenylacrylate), Poly(pentachlorophenyl acrylate), Poly(4-cyanobenzylacrylate), Poly(2-cyanobutyl acrylate), Poly(2-cyanoisobutyl acrylate),Poly(4-cyanobutyl acrylate), Poly(2-cyanoethyl acrylate),Poly(2-cyanoheptyl acrylate), Poly(2-cyanohexyl acrylate),Poly(cyanomethyl acrylate), Poly(2-cyanomethyl acrylate),Poly(5-cyano-3-oxapentyl acrylate), Poly(4-cyanophenyl acrylate),Poly(2-cyanoisopropyl acrylate), Poly(4-cyano-3-thiabutyl acrylate),Poly(6-cyano-3-thiahexyl acrylate), Poly(6-cyano-4-thiahexyl acrylate),Poly(8-cyano-7-thiaoctyl acrylate), Poly(5-cyano-3-thiapentyl acrylate),Poly(cyclododecyl acrylate), Poly(cyclohexyl acrylate),Poly(2-chloroethyl acrylate), Poly[di(chlorodifluoromethyl)fluoromethylacrylate], Poly(2-ethoxycarbonylphenyl acrylate),Poly(3-ethoxycarbonylphenyl acrylate), Poly(4-ethoxycarbonylphenylacrylate), Poly(2-ethoxyethyl acrylate), Poly(3-ethoxypropyl acrylate),Poly(ethyl acrylate), Poly(2-bromoethyl acrylate), Poly(2-ethylbutylacrylate), Poly(2-ethylhexyl acrylate), Poly(ferrocenylethyl acrylate),Poly(ferrocenylmethyl acrylate), Poly(1H,1H-heptafluorobutyl acrylate),Poly(heptafluoroisopropyl acrylate),Poly[5-(heptafluoroisopropoxy)pentyl acrylate],Poly[11-(heptafluoroisopropoxy)undecyl acrylate],Poly[2-(heptafluoropropoxy)ethyl acrylate],Poly[(2-(heptafluorobutoxy)ethyl acrylate],Poly[2-(1,1,2,2-tetrafluoroethoxy)ethyl acrylate],Poly(1H,1H,3H-hexafluorobutyl acrylate), Poly(2,2,2-trifluoroethylacrylate), Poly[2,2-difluoro-2-(2-heptafluorotetrahydrofuranyl)ethylacrylate], Poly(1H,1H-undecafluorohexyl acrylate), Poly(fluoromethylacrylate), Poly(trifluoromethyl acrylate),Poly(1H,1H-pentadecafluorooctyl acrylate),Poly(5,5,6,6,7,7,7-heptafluoro-3-oxaheptyl acrylate),Poly(1H,1H-undecafluoro-4-oxaheptyl acrylate),Poly(1H,1H-nonafluoro-4-oxaheptyl acrylate),Poly(7,7,8,8-tetrafluoro-3,6-dioxaoctyl acrylate),Poly(1H,1H-tridecafluoro-4-oxaoctyl acrylate),Poly(2,2,3,3,5,5,5-heptafluoro-4-oxapentyl acrylate),Poly(4,4,5,5-tetrafluoro-3-oxapentyl acrylate),Poly(5,5,5-trifluoro-3-oxapentyl acrylate), Poly(1H,1H-nonafluoropentylacrylate), Poly(nonafluoroisobutyl acrylate),Poly(1H,1H,5H-octafluoropentyl acrylate), Poly(heptafluoro-2-propylacrylate), Poly[tetrafluoro-3-(heptafluoropropoxy)propyl acrylate],Poly[(tetrafluoro-3-(pentafluoroethoxy)propyl acrylate],Poly[tetrafluoro-3-(trifluoromethoxy)propyl acrylate],Poly(1H,1H-pentafluoropropyl acrylate), Poly(octafluoropentyl acrylate),Poly(heptyl acrylate), Poly(2-heptyl acrylate), Poly(hexadecylacrylate), Poly(hexyl acrylate), Poly(2-ethylhexyl acrylate),Poly(isobornyl acrylate), Poly(isobutyl acrylate), Poly(isopropylacrylate),Poly(1,2:3,4-di-O-isopropylidene-alpha-D-galactopyranos-6-O-ylacrylate), Poly(3-methoxybutyl acrylate), Poly(2-methoxycarbonylphenylacrylate), Poly(3-methoxycarbonylphenyl acrylate),Poly(4-methoxycarbonylphenyl acrylate), Poly(2-methoxyethyl acrylate),Poly(2-ethoxyethyl acrylate), Poly(4-methoxyphenyl acrylate),Poly(3-methoxypropyl acrylate), Poly(3,5-dimethyladamantyl acrylate),Poly(3-dimethylaminophenyl acrylate), Poly(2-methylbutyl acrylate),Poly(3-methylbutyl acrylate), Poly(1,3-dimethylbutyl acrylate),Poly(2-methyl-7-ethyl-4-undecyl acrylate), Poly(2-methylpentylacrylate), Poly(menthyl acrylate), Poly(2-naphthyl acrylate), Poly(nonylacrylate), Poly(octyl acrylate), Poly(2-octyl acrylate), Poly(3-pentylacrylate), Poly(phenethyl acrylate), Poly(phenyl acrylate),Poly(2,4-dinitrophenyl acrylate), Poly(2,4,5-trichlorophenyl acrylate),Poly(2,4,6-tribromophenyl acrylate), Poly(3,4-epoxyhexahydrobenzylacrylate), Poly[alpha-(o-ethyl methylphsphonoxy)-methyl acrylate],Poly(propyl acrylate), Poly(2,3-dibromopropyl acrylate), Poly(tetradecylacrylate), Poly(3-thiabutyl acrylate), Poly(4-thiahexyl acrylate),Poly(5-thiahexyl acrylate, Poly(3-thispentyl acrylate),Poly(4-thiapentyl acrylate), Poly(m-tolyl acrylate), Poly(o-tolylacrylate), Poly(p-tolyl acrylate), Poly(2-ethoxyethyl acrylate),Poly(3-ethoxypropyl acrylate), Poly(cholesteryl acrylate),Poly(2-ethyl-n-hexyl acrylate),Poly(1-oxy-2,2,6,6-tetramethyl-4-piperidyl acrylate),Poly(1,2,2,6,6-pentamethyl-4-piperidyl acrylate),Poly(4-phenylazoxyphenyl acrylate), Poly(ethyl cyanoacrylate),Poly[4-(10,15,20-triphenyl-21H,23H-5-yl)phenyl acrylate],Poly(1,1,5-trihydroperfluoroamyl acrylate), Poly(tributyltin acrylate),Poly(beta-ethoxyethyl acrylate), Poly(3,4-epoxyhexahydrobenzylacrylate), Poly(alpha-chloroacrylnitrile),Poly(alpha-fluoroacrylnitrile), Poly(alpha-methoxy acrylnitrile),Poly(alpha-trifluoromethyl acrylnitrile),Poly(alpha-ethylacrylonitrile), Poly(alpha-isopropylacrylonitrile),Poly(alpha-propylacrylonitrile), Poly(amyl methacrylate),Poly[1-(3-cyanopropyl)acrylonitrile], Poly(t-butyl methacrylate),Poly(hexadecyl methacrylate), Poly(methyl methacrylate),Poly(cyanomethyl methacrylate), Poly(adamantyl methacrylate),Poly(3,5-dimethyladamantyl methacrylate), Poly(benzyl methacrylate),Poly(1-alpha-methylbenzyl methacrylate), Poly(2-bromoethylmethacrylate), Poly(2-t-butylaminoethyl methacrylate), Poly(butylmethacrylate), Poly(sec-butyl methacrylate), Poly(tert-butylmethacrylate), Poly(ethylbutyl metbacrylate),Poly(4-phenylbutyl-1-methacrylate), Poly(2-phenylethyl-1-methacrylate),Poly(cetyl methacrylate), Poly(p-cetyloxybenzoyl methacrylate),Poly(2-chloroethyl methacrylate), Poly(cyanomethyl methacrylate),Poly(2-cyanoethyl methacrylate), Poly(4-cyanomethylphenyl methacrylate),Poly(4-cyanophenyl methacrylate), Poly(cyclohexyl methacrylate),Poly(p-t-butylcyclohexyl methacrylate), Poly(4-t-butylcyclohexylmethacrylate), Poly(cyclobutyl methacrylate), Poly(cyclobutylmethylmethacrylate), Poly(cyclododecyl methacrylate), Poly(2-cyclohexylethylmethacrylate), Poly(cyclohexylmethyl methacrylate), Poly(cyclopentylmethacrylate), Poly(cyclooctyl methacrylate), Poly(decyl methacrylate),Poly(n-decyl methacrylate), Poly(dodecyl methacrylate), Poly(n-decosylmethacrylate), Poly(diethylaminoethyl methacrylate),Poly(dimethylaminoethyl methacrylate), Poly(2-ethylhexyl methacrylate),Poly(ethyl methacrylate), Poly(acetoxyethyl methacrylate),Poly(2-methoxyethyl methacrylate), Poly(2-ethylsulfinylethylmethacrylate), Poly(ferrocenylethyl methacrylate), Poly(ferrocenylmethylmethacrylate), Poly(N-methyl-N-phenyl-2-aminoethyl methacrylate),Poly(2-N,N-dimethylcarbamoyloxyethyl methacrylate), Poly(2-acetoxymethacrylate), Poly(2-bromoethyl methacrylate), Poly(2-chloroethylmethacrylate), Poly(1H,1H-heptafluorobutyl methacrylate),Poly(1H,1H,7H-dodecafluoroheptyl methacrylate),Poly(1H,1H,9H-hexadecafluorononyl methacrylate),Poly(1H,1H,5H-octafluoropentyl methacrylate),Poly(1,1,1-trifluoro-2-propyl methacrylate), Poly(trifluoroisopropylmethacrylate), Poly(hexadecyl methacrylate), Poly(hexyl methacrylate),Poly(isobornyl methacrylate), Poly(isobutyl methacrylate),Poly(isopropyl methacrylate),Poly(1,2:3,4-di-O-isopropylidene-alpha-D-galactopyranos-6-O-ylmethacrylate), Poly(2,3-O-isopropylidene-DL-glyceritol-1-O-ylmethacrylate), Poly(nonyl methacrylate), Poly(methacrylic acidanhydride), Poly(4-methoxycarbonylphenyl methacrylate),Poly(3,5-dimethyladamantyl methacrylate), Poly(dimethylaminoethylmethacrylate), Poly(2-methylbutyl methacrylate), Poly(1,3-dimethylbutylmethacrylate), Poly(3,3-dimethylbutyl methacrylate),Poly(3,3-dimethyl-2-butyl methacrylate), Poly(3,5,5-trimethylhexylmethacrylate), Poly(trimethylsilyl methacrylate),Poly[(2-nitratoethyl)methacrylate], Poly(octadecyl methacrylate),Poly(octyl methacrylate), Poly(n-octadecyl methacrylate),Poly(3-oxabutyl methacrylate), Poly(pentyl methacrylate), Poly(neopentylmethacrylate), Poly(phenethyl methacrylate), Poly(phenyl methacrylate),Poly(2,6-diisopropylphenyl methacrylate), Poly(2,6-dimethylphenylmethacrylate), Poly(2,4-dinitrophenyl methacrylate), Poly(diphenylmethylmethacrylate), Poly(4-t-butylphenyl methacrylate), Poly(2-t-butylphenylmethacrylate), Poly(o-ethylphenyl methacrylate), Poly(p-ethylphenylmethacrylate), Poly(m-chlorophenyl methacrylate), Poly(m-nitrophenylmethacrylate), Poly(propyl methacrylate), Poly(tetradecyl methacrylate),Poly(butyl butoxycarbonyl methacrylate), Poly(tetradecyl methacrylate),Poly(ethylidene dimethacrylate), Poly(3,3,5-trimethylcyclohexylmethacrylate), Poly(2-nitro-2-methylpropyl methacrylate),Poly(triethylcarbinyl methacrylate), Poly(triphenylmethyl methacrylate),Poly(1,1-diethylpropyl methacrylate), Poly(ethyl glycolatemethacrylate), Poly(3-methylcyclohexyl methacrylate),Poly(4-methylcyclohexyl metbacrylate), Poly(2-methylcyclohexylmethacrylate), Poly(1-methylcyclohexyl methacrylate), Poly(bornylmethacrylate), Poly(tetrahydrofurfuryl methacrylate), Poly(vinylmethacrylate), Poly(2-chloroethyl methacrylate),Poly(2-diethylaminoethyl methacrylate), Poly(2-chlorocyclohexylmethacrylate), Poly(2-aminoethyl methacrylate), Poly(furfurylmethacrylate), Poly(methylmercaptyl methacrylate),Poly(2,3-epithiopropyl methacrylate), Poly(ferrocenylethylmethacrylate), Poly[2-(N,N-dimethylcarbamoyloxy)ethyl methacrylate],Poly(butyl butoxycarbonyl methacrylate), Poly(cyclohexylchloroacrylate), Poly(ethyl chloroacrylate), Poly(ethyl ethoxycarbonylmethacrylate), Poly(ethyl ethacrylate), Poly(ethyl fluoromethacrylate),Poly(hexyl hexyloxycarbonyl methacrylate),Poly(1,1-dihydropentadecafluorooctyl methacrylate),Poly(heptafluoroisopropyl methacrylate), Poly(heptadecafluorooctylmethacrylate), Poly(1-hydrotetrafluoroethyl methacrylate),Poly(1,1-dihydrotetrafluoroisopropyl methacrylate),Poly(1-hydrohexafluorobutyl methacrylate), Poly(1-nonafluorobutylmethacrylate), Poly(1,3-dichloropropyl methacrylate),Poly[2-chloro-1-(chloromethyl)ethyl methacrylate], Poly(butylmercaptylmethacrylate), Poly(1-phenyl-n-amyl methacrylate),Poly[2-heptoxycarbonyl-1-heptoxycarbonylethylene)ethylene],Poly(2-t-butylphenyl methacrylate), Poly(4-cetyloxycarbonylphenylmethacrylate), Poly(1-phenylethyl methacrylate), Poly(p-methoxybenzylmethacrylate), Poly(1-phenylallyl methacrylate), Poly(p-cyclohexylphenylmethacrylate), Poly(2-phenylethyl methacrylate),Poly[1-(chlorophenyl)cyclohexyl methacrylate], Poly(1-phenylcyclohexylmethacrylate), Poly[2-(phenylsulfonyl)ethyl methacrylate], Poly(m-cresylmethacrylate), Poly(o-cresyl methacrylate), Poly(2,3-dibromopropylmethacrylate), Poly(1,2-diphenylethyl methacrylate), Poly(o-chlorobenzylmethacrylate), Poly(m-nitrobenzyl methacrylate), Poly(2-diphenylmethacrylate), Poly(4-diphenyl methacrylate), Poly(alpha-naphthylmethacrylate), Poly(beta-naphthyl methacrylate), Poly(alpha-naphthylcarbinyl methacrylate), Poly(2-ethoxyethyl methacrylate), Poly(laurylmethacrylate), Poly(pentabromophenyl methacrylate), Poly(o-bromobenzylmethacrylate), Poly(o-chlorodiphenylmethyl methacrylate),Poly(pentachlorophenyl methacrylate), Poly(2-diethylamino methacrylate),Poly(2-fluoroethyl mathacrylate), Poly(hexadecyl methacrylate),Poly(2-ethylbutyl methacrylate),Poly[4-(4-hexadecyloxy-benzoyloxy)phenyl methacrylate],Poly(D,L-diisobornyl methacrylate), Poly(decahydro-beta-naphthylmethacrylate), Poly(5-p-menthyl methacrylate), Poly(methyl butacrylate),Poly(methyl ethacrylate), Poly[(2-methylsulfinyl)ethylacrylate],Poly(methylphenylacrylate), Poly[4-(4-nonyloxy-benzoyloxy)-phenylmethacrylate], Poly(tetrahydrofurfuryl methacrylate),Poly[2-(triphenylmethoxy)ethyl methacrylate], Poly(cetyl methacrylate),Poly(2,3-epoxypropyl methacrylate), Poly(pentachlorophenylmethacrylate), Poly(pentafluorophenyl methacrylate),Poly[6-(anisyloxycarbonylphenoxy)hexyl methacrylate],Poly(ethyl-alpha-bromoacrylate),Poly[1-(2-N-cyclohexyl-N-methyl-carbamoyloxy)ethyl methacrylate],Poly[1-(2-N,N-diethylcarbamoyloxy)ethyl methacrylate],Poly[(2-N,N-diethylcarbamoyloxy)-2-methylethyl methacrylate],Poly(n-docosyl methacrylate), Poly(2,5-dimethylpyrozolyl methacrylate),Poly[11-(hexadecyl-dimethylammonio)-undecyl methacrylate],Poly[2-(4-methyl-1-piperazinylcarbonyloxy)ethyl methacrylate],Poly[(2-morpholino-carbonyloxy)ethylmethacrylate],Poly[1-(1-nonyloxy-4-phenoxycarbonyl)phenyl methacrylate],Poly(1,2,2,6,6-pentamethyl-4-piperidyl methacrylate),Poly(propionyloxyethyl methacrylate),Poly[3-(8-oxy]-7,7,9,9-tetramethyl-2,4-dioxo-1,3,8-triazaspiro(4,5)-dec-3-yl)propylmethacrylate], Poly(n-stearyl methacrylate),Poly[4-(1,1,3,3-tetramethylbutyl)phenyl methacrylate], Poly(o-tolylmethacrylate), Poly(p-tolyl methacrylate), Poly(2,4,5-trichlorophenylmethacrylate), Poly(n-tridecyl methacrylate), Poly(triphenylmethylmethacrylate), Poly(trityl methacrylate),Poly(tetrahydro-4H-pyranyl-2-methacrylate), Poly(tridecyl methacrylate),Poly[2-(triphenylmethoxy)ethyl methacrylate],Poly[2-(4-methyl-1-piperazinylcarbonyloxy)-2-methylethyl methacrylate],Poly(p-methoxyphenyl-oxycarbonyl-p-phenoxyhexamethylene methacrylate),Poly(diphenyl-2-pyridylmethyl methacrylate),Poly(diphenyl-4-pyridylmethyl methacrylate), Poly(triphenylmethylmethacrylate), Poly(hexyleneoxyphenylenecarboxyphenyleneoxymethylenemethacrylate), Poly[4-(1,1,3,3-tetramethylbutyl)phenylmethacrylate],Poly(glycidyl methacrylate), Poly(2,2,6,6-tetramethyl-4-piperidinylmethacrylate), Poly[(2,2-dimethyl-1,3-dioxolane-4-yl)methylmethacrylate], Poly(alpha-alpha-dimethylbenzyl methacrylate),Poly(1,1-diphenylethyl methacrylate), Poly(2,3-epithiopropylmethacrylate), Poly(dicyclopentadienyltitanate dimethacrylate),Poly(diethylaminoethyl methacrylate), Poly(5-oxo-pyrrolidinylmethylmethacrylate), Poly(ethyl-alpha-bromoacrylate),Poly(isopropyl-alpha-bromoacrylate), Poly(methyl-alpha-bromoacrylate),Poly(n-pentyl-alpha-bromoacrylate), Poly(n-propyl-alpha-bromoacrylate),Poly(methyl alpha-trifluoromethylacrylate), Poly(phenylalpha-bromoacrylate), Poly(sec-butyl-alpha-bromoacrylate),Poly(cyclohexyl-alpha-bromoacrylate),Poly(methyl-alpha-bromomethacrylate), Poly(butyl chloroacrylate),Poly(sec-butyl chloroacrylate), Poly(methyl chloroacrylate),Poly(isobutyl chloroacrylate), Poly(isopropyl chloroacrylate),Poly(cyclohexyl chloroacrylate), Poly(2-chloroethyl chloroacrylate),Poly[1-methoxycarbonyl-1-methoxycarbonylmethylene)ethylene], Poly(methylchloroacrylate), Poly(ethyl alpha-chloroacrylate), Poly(methylbeta-chloroacrylate), Poly(cyclohexyl alpha-ethoxyacrylate), Poly(methylfluoroacrylate), Poly(methyl fluoromethacrylate), Poly(methylphenylacrylate), Poly(propyl chloroacrylate), Poly(methylcyanoacrylate), Poly(ethyl cyanoacrylate), Poly(butylcyanoacrylate),Poly(sec-butyl thiolacrylate), Poly(isobutyl thiolacrylate), Poly(ethylthioacrylate), Poly(methyl thioacrylate), Poly(butyl thioacrylate),Poly(isopropyl thiolacrylate), Poly(propyl thiolacrylate), Poly(phenylthiomethacrylate), Poly(cyclohexyl thiomethacrylate),Poly(o-methylphenylthio methacrylate),Poly(nonyloxy-1,4-phenyleneoxycarbonylphenyl methacrylate),Poly(4-methyl-2-N,N-dimethyl aminopentyl methacrylate),Poly[alpha-(4-chlorobenzyl)ethyl acrylate],Poly[alpha-(4-cyanobenzyl)ethyl acrylate],Poly[alpha-(4-methoxybenzyl)ethyl acrylate], Poly(alpha-acetoxy ethylacrylate), Poly(ethyl alpha-benzylacrylate), Poly(methylalpha-benzylacrylate), Poly(methyl alpha-hexylacrylate), Poly(ethylalpha-fluoroacrylate), Poly(methyl alpha-fluoroacrylate), Poly(methylalpha-isobutylacrylate), Poly(methyl alpha-isopropylacrylate),Poly(methyl alpha-methoxyacrylate), Poly(butyl alpha-phenylacrylate),Poly(chloroethyl alpha-phenylacrylate), Poly(methylalpha-phenylacrylate), Poly(propyl alpha-phenylacrylate), Poly(methylalpha-propylacrylate), Poly(methyl alpha-sec-butylacrylate), Poly(methylalpha-trifluoromethylacrylate), Poly(ethyl alpha-acetoxyacrylate),Poly(ethyl beta-ethoxyacrylate), Poly(methacryloyl chloride),Poly(methacryloylactone), Poly(meethylenebutyrolactone),Poly(acryloylpyrrolidone), Poly[butylN-(4-carbethoxyphenyl)itaconamate], Poly[ethylN-(4-carbethoxyphenyl)itaconamate], Poly[methylN-(4-carbethoxyphenyl)itaconamate], Poly[propylN-(4-carbethoxyphenyl)itaconamate], Poly[ethylN-(4-chlorophenyl)itaconamate], Poly[methylN-(4-chlorophenyl)itaconamate], Poly[propylN-(4-chlorophenyl)itaconamate], Poly[butylN-(4-methoxyphenyl)itaconamate], Poly[ethylN-(4-methoxyphenyl)itaconamate], Poly[methylN-(4-methoxyphenyl)itaconamate], Poly[propylN-(4-methoxyphenyl)itaconamate], Poly[butylN-(4-methylphenyl)itaconamate], Poly[ethylN-(4-methylphenyl)itaconamate], Poly[methylN-(4-methylphenyl)itaconamate], Poly[propylN-(4-methylphenyl)itaconamate], Poly[butyl N-phenyl itaconamate],Poly[ethyl N-phenyl itaconamate], Poly[methyl N-phenyl itaconamate],Poly[propyl N-phenyl itaconamate], Poly(diamyl itaconate), Poly(dibutylitaconate), Poly(diethyl itaconate), Poly(dioctyl itaconate),Poly(dipropyl itaconate), Polystyrene, Poly[(p-t-butyl)-styrene],Poly[(o-fluoro)-styrene], Poly[(p-fluoro)-styrene],Poly[(alpha-methyl)-styrene], Poly[(alpha-methyl)(p-methyl)-styrene],Poly[(m-methyl)-styrene], Poly[(o-methyl)-styrene],Poly[(o-methyl)(p-fluoro)-styrene], Poly[(p-methyl)-styrene],Poly(trimethylsilylstyrene), Poly(beta-nitrostyrene),Poly(4-acetylstyrene), Poly(4-acetoxystyrene), Poly(4-p-anisoylstyrene),Poly(4-benzoylstyrene), Poly[(2-benzoyloxymethyl)styrene],Poly[(3-(4-biphenylyl)styrene], Poly[(4-(4-biphenylyl)styrene],Poly(5-bromo-2-butoxystyrene), Poly(5-bromo-2-ethoxystyrene),Poly(5-bromo-2-isopentyloxystyrene), Poly(5-bromo-2-isopropoxystyrene),Poly(4-bromostyrene), Poly(2-butoxycarbonylstyrene),Poly(4-butoxycarbonylstyrene), Poly(4-[(2-butoxyethoxy)methyl]styrene),Poly(2-butoxymethylstyrene), Poly(4-butoxymethylstyrene),Poly[4-(sec-butoxymethyl)styrene], Poly(4-butoxystyrene),Poly(5-t-butyl-2-methylstyrene), Poly(4-butylstyrene),Poly(4-sec-butylstyrene), Poly(4-t-butylstyrene),Poly(4-butyrylstyrene), Poly(4-chloro-3-fluorostyrene),Poly(4-chloro-2-methylstyrene), Poly(4-chloro-3-methylstyrene),Poly(2-chlorostyrene), Poly(3-chlorostyrene), Poly(4-chlorostyrene),Poly(2,4-dichlorostyrene), Poly(2,5-dichlorostyrene),Poly(2,6-dichlorostyrene), Poly(3,4-dichlorostyrene),Poly(2-bromo-4-trifluoromethylstyrene), Poly(4-cyanostyrene),Poly(4-decylstyrene), Poly(4-dodecylstyrene),Poly(2-ethoxycarbonylstyrene), Poly(4-ethoxycarbonylstyrene),Poly[4-(2-ethoxymethyl)styrene], Poly(2-ethoxymethylstyrene),Poly(4-ethoxystyrene), Poly[4-(2-diethylaminoethoxycarbonyl)styrene],Poly(4-diethylcarbamoylstyrene), Poly[4-(1-ethylhexyloxymethyl)styrene],Poly(2-ethylstyrene), Poly(3-ethylstyrene), Poly(4-ethyl styrene),Poly[4-(pentadecafluoroheptyl)styrene], Poly(2-fluoro-5-methylstyrene),Poly(4-fluorostyrene), Poly(3-fluorostyrene),Poly(4-fluoro-2-trifluoromethyl styrene), Poly(p-fluoromethyl styrene),Poly(2,5-difluorostyrene), Poly(2,3,4,5,6,-pentafluorostyrene),Poly(perfluorostyrene), Poly(alpha,beta,beta-trifluorostyrene),Poly(4-hexadecylstyrene), Poly(4-hexanoylstyrene),Poly(2-hexyloxycarbonylstyrene), Poly(4-hexyloxycarbonylstyrene),Poly(4-hexyloxymethylstyrene), Poly(4-hexylstyrene),Poly(4-iodostyrene), Poly(2-isobutoxycarbonylstyrene),Poly(4-isobutoxycarbonylstyrene), Poly(2-isopentyloxycarbonylstyrene),Poly(2-isopentyloxymethylstyrene), Poly(4-isopentyloxystyrene),Poly(2-isopropoxycarbonylstyrene), Poly(4-isopropoxycarbonylstyrene),Poly(2-isopropoxymethylstyrene), Poly(4-isopropylstyrene),Poly(4-isopropyl-alpha-methylstyrene),Poly(4-trimethylsilyl-alpha-methylstyrene),Poly(2,4-diisopropylstyrene), Poly(2,5-diisopropylstyrene),Poly(beta-methylstyrene), Poly(2-methoxymethylstyrene),Poly(2-methoxycarbonylstyrene), Poly(4-methoxycarbonylstyrene),Poly(4-methoxymethylstyrene), Poly(4-methoxy-2-methylstyrene),Poly(2-methoxystyrene), Poly(4-methoxystyrene), Poly(4-N,N-dimethylaminostyrene), Poly(2-methylaminocarbonylstyrene),Poly(2-dimethylaminocarbonylstyrene),Poly(4-dimethylaminocarbonylstyrene),Poly[2-(2-dimethylaminoethoxycarbonyl)styrene],Poly[4-(2-dimethylaminoethoxycarbonyl)styrene], Poly(2-methylstyrene),Poly(3-methylstyrene), Poly(4-methylstyrene), Poly(4-methoxystyrene),Poly(2,4-dimethylstyrene), Poly(2,5-dimethylstyrene),Poly(3,4-dimethylstyrene), Poly(3,5-dimethylstyrene),Poly(2,4,5-trimethylstyrene), Poly(2,4,6-trimethylstyrene),Poly(3-[bis(trimethylsiloxy)boryl]styrene),Poly(4-[bis(trimethylsiloxy)boryl]styrene),Poly(4-morpholinocarbonylstyrene),Poly[4-(3-morpholinopropionyl)styrene], Poly(4-nonadecylstyrene),Poly(4-nonylstyrene), Poly(4-octadecylstyrene), Poly(4-octanoylstyrene),Poly[4-(octyloxymethyl)styrene], Poly(2-octyloxystyrene),Poly(4-octyloxystyrene), Poly(2-pentyloxycarbonylstyrene),Poly(2-pentyloxymethylstyrene), Poly(2-phenethyloxymethylstyrene),Poly(2-phenoxycarbonylstyrene), Poly(4-phenoxystyrene),Poly(4-phenylacetylstyrene), Poly(2-phenylaminocarbonylstyrene),Poly(4-phenylstyrene), Poly(4-piperidinocarbonylstyrene),Poly[4-(3-piperidinopropionyl)styrene], Poly(4-propionylstyrene),Poly(2-propoxycarbonylstyrene), Poly(4-propoxycarbonylstyrene),Poly(2-propoxymethylstyrene), Poly(4-propoxymethylstyrene),Poly(4-propoxystyrene), Poly(4-propoxysulfonylstyrene),Poly(4-tetradecylstyrene), Poly(4-p-toluoylstyrene),Poly(4-trimethylsilylstyrene), Poly[2-(2-thio-3-methylpentyl)styrene],Poly[9-(2-methylbutyl)-2-vinyl carbazole],Poly[9-(2-methylbutyl)-3-vinyl carbazole], Poly(3-sec-butyl-9-vinylcarbazole), Poly[p-(p-tolylsulfinyl)styrene], Poly(4-valerylstyrene),Poly[(4-t-butyl-dimethylsilyl)oxy styrene], Poly(4-isopropyl-2-methylstyrene), Poly[1-(4-formylphenyl)ethylene], Poly(alpha-methoxystyrene),Poly(alpha-methylstyrene), Poly(p-octylamine sulfonate styrene),Poly(m-divinylbenzene), Poly(p-divinylbenzene), Polybutadiene(1,4-addition), Polybutadiene (1,2-addition),(2-t-butyl)-cis-1,4-poly-1,3-butadiene,(2-chloro)-trans-1,4-poly-1,3-butadiene,(2-chloro)-cis-1,4-poly-1,3-butadiene,(1-cyano)-trans-1,4-poly-1,3-butadiene,(1-methoxy)-trans-1,4-poly-1,3-butadiene,(2,3-dichloro)-trans-1,4-poly-1,3-butadiene,(2,3-dimethyl)-trans-1,4-poly-1,3-butadiene,(2,3-dimethyl)-cis-1,4-poly-1,3-butadiene,(2-methyl)-cis-1,4-poly-1,3-butadiene,(2-methyl)-trans-1,4-poly-1,3-butadiene,(2-methyl-3-chloro)-trans-1,4-poly-1,3-butadiene,(2-methylacetoxy)-trans-1,4-poly-1,3-butadiene,(2-propyl)-trans-1,4-poly-1,3-butadiene, Poly(2-decyl-1,3-butadiene),Poly(2-heptyl-1,3-butadiene), Poly(2-isopropyl-1,3-butadiene),Poly(2-t-butyl-1,3-butadiene),[1,4-(4,4′-diphenyleneglutarate)]-1,4-poly-1,3-butadiene,Poly(2-chloromethyl-1,3-butadiene),Poly(ethyl-1-carboxylate-1,3-butadiene),Poly(1-diethylamino-1,3-butadiene), Poly(diethyl1,4-carboxylate-1,3-butadiene), Poly(1-acetoxy-1,3-butadiene),Poly(1-ethoxy-1,3-butadiene), Poly(2-phthalidomethyl-1,3-butadiene),Poly(2,3-bis(diethylphosphono-1,3-butadiene),Poly(hexafluoro-1,3-butadiene), Poly(2-fluoro-1,3-butadiene),Poly(1-phthalimido-1,3-butadiene), Poly(1,4-poly-1,3-cyclohexylene),1,12-poly-1,11-dodecadiyne, 1,2-poly-1,3-pentadiene,(4-methyl)-1,2-poly-1,4-pentadiene, Poly(perfluoro-1,4-pentadiene),Poly(1-ferrocenyl-1,3-butadiene), Poly(perfluorobutadiene),Poly(1-phenylbutadiene), Poly(spiro-2,4-hepta-4,6-diene),Poly(1,1,2-trichlorobutadiene), Poly(1,3-pentadiene),1,4-poly-1,3-heptadiene, (6-methyl)-trans-1,4-poly-1,3-heptadiene,(5-methyl)-trans-1,4-poly-1,3-heptadiene,(3,5-dimethyl)-1,4-poly-1,3-heptadiene,(6-phenyl)-1,4-poly-1,3-heptadiene, 1,4-poly-trans-1,3-hexadiene,(5-methyl)-trans-1,4-poly-1,3-hexadiene,(5-phenyl)-trans-1,4-poly-1,3-hexadiene, trans-2,5-poly-2,4-hexadiene,(2,5-dimethyl)-trans-2,5-poly-2,4-hexadiene, Poly(1,5-hexadiene),1,4-poly-1,3-octadiene, 1,4-poly-chloroprene, 1,4-poly-isoprene,Poly(hexatriene), Poly(trichlorohexatriene), 2,5-poly-2,4-hexadienoicacid, diisopropyl ester, 2,5-poly-2,4-hexadienoic acid, butyl ester,2,5-poly-2,4-hexadienoic acid, ethyl ester, 2,5-poly-2,4-hexadienoicacid, isoamyl ester, 2,5-poly-2,4-hexadienoic acid, isobutyl ester,2,5-poly-2,4-hexadienoicacid, isopropyl ester, 2,5-poly-2,4-hexadienoicacid, methyl ester, 2,5-poly-2,4-hexadiyne,[1,6-di(N-carbazoyl))-2,5-poly-2,4-hexadiyne, 1,9-poly-1,8-nonadiyne,1,4-poly-1,3-octadene, 1,2-poly-1,3-pentadiene,(4-methyl)-1,2-poly-1,3-pentadiene, 1,4-poly-1,3-pentadiene,(2-methyl)-1,4-poly-1,3-pentadiene, 2,5-poly-5-phenyl-2,4-pentadienoicacid, butyl ester, 2,5-poly-5-phenyl-2,4-pentadienoic acid, methylester, Poly(4-trans-4-ethoxy-2,4-pentadienoate),Poly(trans-4-ethoxy-2,4-pentadienonitrile),1,24-poly-1,11,13,23-tetracisatetrayne, Poly(3-hydroxybutyric acid),Poly(10-hydroxycapric acid), Poly(3-hydroxy-3-trichloromethyl-propionicacid), Poly(2-hydroxyacetic acid), Poly(dimethyl-2-hydroxyacetic acid),Poly(diethyl-2-hydroxyacetic acid), Poly(isopropyl-2-hydroxyaceticacid), Poly(3-hydroxy-3-butenoic acid), Poly(6-hydroxy-carproic acid),Poly(5-hydroxy-2-(1,3-dioxane)-carprylic acid], Poly(7-hydroxynanthicacid), Poly[(4-methyl)-7-hydroxynanthic acid],Poly[4-hydroxymethylene-2-(1,3-dioxane)-carprylic acid],Poly(5-hydroxy-3-oxavaleric acid),Poly(2,3,4-trimethoxy-5-hydroxyvaleric acid), Poly(2-hydroxypropionicacid), Poly(3-hydroxypropionic acid),Poly(2,2-bischloromethyl-3-hydroxypropionic acid),Poly(3-chloromethyl-3-hydroxypropionic acid),Poly(2,2-butyl-3-hydroxypropionic acid),Poly(3-dichloromethyl-3-hydroxypropionic acid),Poly(2,2-diethyl-3-hydroxypropionic acid),Poly(2,2-dimethyl-3-hydroxypropionic acid),Poly(3-ethyl-3-hydroxypropionic acid),Poly(2-ethyl-2-methyl-3-hydroxypropionic acid),Poly(2-ethyl-2-methyl-1,1-dichloro-3-hydroxypropionic acid),Poly(3-isopropyl-3-hydroxypropionic acid),Poly(2-methyl-3-hydroxypropionic acid), Poly(3-methyl-3-hydroxypropionicacid), Poly(2-methyl-2-propyl-3-hydroxypropionic acid),Poly(3-trichloromethyl-3-hydroxypropionic acid),Poly(carbonoxide-alt-ethylene),Poly(oxycarbonyl-1,5-dimethylpentamethylene),Poly(oxycarbonylethylidene), Poly(oxycarbonylisobutylidene),Poly(oxycarbonylisopentylidene), Poly(oxycarbonylpentamethylene),Poly(oxycrabonyl-3-methylhexamethylene),Poly(oxycarbonyl-2-methylpentamethylene),Poly(oxycarbonyl-3-methylpentamethylene),Poly(oxycarbonyl-4-methylpentamethylene),Poly(oxycarbonyl-1,2,3-trimethyloxytetramethylene),Poly(2-mercaptocarproic acid), Poly(4-methyl-2-mercaptocarproic acid),Poly(2-mercaptoacetic acid), Poly(2-methyl-2-mercaptoacetic acid),Poly(3-mercaptopropionoic acid), Poly(2-phthalimido-3-mercaptopropionoicacid), Poly[2-(p-toluenesulfonamido)-3-mercaptopropionic acid],Poly(thiodipropionic anhydride), Poly(ethyl alpha-cyanocinnamate),Poly(cinnamonitrile), Poly(alpha-cyanocinnamonitrile), Poly(N-methylcitraconimide), Poly(methyl alpha-acetyl crotonate), Poly(ethylalpha-carbethoxy crotonate), Poly(ethyl alpha-chlorocrotonate),Poly(ethyl alpha-cyanocrotonate), Poly(methyl alpha-methoxycrotonate),Poly(methyl alpha-methylcrotonate), Poly(ethyl crotonate), Poly(diethylfumarate), Poly(vinyl acetalacetate), Poly(vinyl chloroacetate),Poly(vinyl dichloroacetate), Poly(vinyl trichloroacetate),Poly(trifluorovinyl acetate), Poly(propenyl acetate),Poly(2-chloropropenyl acetate), Poly(2-methylpropenyl acetate),Poly(vinyl chloroacetate), Poly(vinyl benzoate), Poly(p-t-butylvinylbenzoate), Poly(vinyl 4-chlorobenzoate), Poly(vinyl3-trimethylsilylbenzoate), Poly(vinyl 4-trimethylsilylbenzoate),Poly(p-acryloyloxyphenyl benzoate), Poly(vinyl butyrate), Poly(vinyl1,2-phenylbutyrate), Poly(vinyl caproate), Poly(vinyl cinnamate),Poly(vinyl decanoate), Poly(vinyl dodecanoate), Poly(vinylformate),Poly(methyl allyl fumarate), Poly(vinyl hexanoate), Poly(vinyl2-ethylhexanoate), Poly(vinyl hexadeconoate), Poly(vinyl isobutyrate),Poly(vinyl isocaproate), Poly(vinyl laurate), Poly(vinyl myristate),Poly(vinyl octanoate), Poly(methyl allyl oxalate), Poly(octyl allyloxalate), Poly(1-vinyl-palmitate), Poly(t-butyl-4-vinyl perbenzoate),Poly(vinyl propionoate), Poly(vinyl pivalate), Poly(vinyl stearate),Poly(2-chloropropenyl acetate), Poly(vinyl hendecanoate), Poly(vinylthioacetate), Poly(vinylhydroquinone dibenzoate), Poly(vinylisocyanate), Poly(N-vinyl-ethyl carbamate), Poly(N-vinyl-t-butylcarbamate), Poly(N,N-diethyl vinyl carbamate), Poly(2-chloro-propenylacetate), Poly(vinylhydroquinone dibenzoate), Poly(ethyltrans-4-ethoxy-2,4-pentadienoate), Poly(triallyl citrate), Poly(vinyl12-ketostearate), Poly(vinyl 2-ethylhexanoate), Poly(vinylenecarbonate), Poly(divinyl adipate), Poly(vinyl hexadecanoate), Poly(vinylpelargonate), Poly(vinyl thioisocyanate), Poly(vinyl valerate),Poly(diallyl-beta-cyanoethylisocyanurate), Poly(diallylcyanamide),Poly(triallyl citrate), Poly(triallyl cyanurate), Poly(triallylisocyanurate), Poly[3-(1-cyclohexenyl)isopropenyl acetate),Poly(isopropenyl acetate), Poly(isopropenylisocyanate), Poly(vinyldiethyl phosphate), Poly(allyl acetate), Poly(vinyl phenylisocyanate),Poly(benzylvinylether), Poly(butylvinylether),Poly(2-methylbutylvinylether), Poly(sec-butylvinylether),Poly(1-methyl-sec-butylvinylether), Poly(t-butylvinylether),Poly(butylthioethylene), Poly(1-butoxy-2-chloroethylene),cis,Poly(1-butoxy-2-chloroethylene),trans,Poly(1-chloro-2-isobutoxyethylene),trans,Poly(1-isobutoxy-2-methylethylene),trans, Poly(ethylvinyl ether),Poly(2-chloroethylvinyl ether), Poly(2-bromoethylvinyl ether),Poly(vinylbutyl sulfonate), Poly(2-methoxyethylvinyl ether),Poly(2,2,2-trifluoroethylvinyl ether), Poly(isobutylvinylether),Poly(isopropylvinylether), Poly(methylvinylether), Poly(octylvinylether), Poly(alpha-methylvinylether), Poly(n-pentylvinylether),Poly(propylvinylether), Poly(1-methylpropylvinylether), Poly(decylvinylether), Poly(dodecylvinyl ether), Poly(isobutylpropenyl ether),Poly(cyclohexyloxyethylene), Poly(hexadecyloxyethylene),Poly(octadecyloxyethylene), Poly(1-bornyloxyethylene),Poly(1-cholesteryloxyethylene),Poly(1,2-5,6-diisopropylidene-alpha-D-glucofuranosyl-3-oxyethylene),Poly(1-menthyloxyethylene), Poly(1-alpha-methylbenzyloxyethylene),Poly[3-beta-(styryloxy)methane], Poly(2-phenylvinyl 2-methylbutylether), Poly(2-phenylvinyl 3-methylpentyl ether),Poly[(2-ethylhexyloxy)ethylene], Poly(ethylthioethylene),Poly(dodecafluorobutoxyethylene),Poly(2,2,2-trifluoroethoxytrifluoroethylene),Poly[1,1-bis(trifluoromethoxy)difluoroethylene],Poly(1,1-difluoro-2-trifluoromethoxymethylene),Poly(1,2-difluoro-1-trifluoromethoxymethylene),Poly(hexafluoromethoxyethylene), Poly[(heptafluoro-2-propoxy)ethylene],Poly(hexyloxyethylene), Poly(isobutoxyethylene), Poly(isopropenyl methylether), Poly(isopropoxyethylene), Poly(methoxyethylene),Poly(2-methoxypropylene), Poly(2,2-dimethylbutoxyethylene),Poly(methylthioethylene), Poly(neopentyloxyethylene),Poly(octyloxyethylene), Poly(pentyloxyethylene), Poly(propoxyethylene),Poly(1-acetyl-1-fluoroethylene), Poly(4-bromo-3-methoxybenzoylethylene),Poly(4-t-butylbenzoylethylene), Poly(4-chlorobenzoylethylene),Poly(4-ethylbenzoylethylene), Poly(4-isopropylbenzoylethylene),Poly(4-methoxybenzoylethylene), Poly(3,4-dimethylbenzoylethylene),Poly(4-propylbenzoylethylene), Poly(p-toluoylethylene), Poly(vinylisobutyl sulfide), Poly(vinyl methyl sulfide), Poly(vinyl phenylsulfide), Poly(vinyl ethyl sulfoxide), Poly(vinyl ethyl sulfide),Poly(t-butyl vinyl ketone), Poly(isopropenyl methyl ketone), Poly(methylvinyl ketone), Poly(phenyl vinyl ketone), Poly(2-methylbutyl vinylketone), Poly(3-methylpentyl vinyl ketone), Poly(isopropenylisocyanate),Poly(vinyl chloromethyl ketone), Poly(vinyl 2-chlorocyclohexyl ketone),Poly(vinyl 4-chlorocyclohexyl ketone), Poly(2-chloroacetaldehyde),Poly(2,2-dichloroacetaldehyde), Poly(2,2,2-trichloroacetaldehyde),Poly(2-butene oxide), Poly(2-methyl-2-butene oxide), Poly(butadieneoxide), Poly(butyraldehyde), Poly(crotonaldehyde), Poly(valeraldehyde),Poly(1,3-cyclobutyleneoxymethylene oxide),Poly[(2,2,4,4-tetramethyl)-1,3-cyclobutyleneoxymethylene oxide],Poly(decamethylene oxide), Poly(dodecamethylene oxide), Poly(ethylenetrimethylene oxide), Poly(1,1-bischloromethyl-ethylene oxide),Poly(bromomethyl-ethyleneoxide), Poly(t-butyl-ethyleneoxide),Poly(chloromethyl-ethylene oxide), Poly(1,2-dichloromethyl-ethyleneoxide), Poly(1-fluoroethylene oxide), Poly(isopropyl-ethyleneoxide),Poly(neopentyl-ethyleneoxide), Poly(tetrafluoro-ethylene oxide),Poly(tetramethyl-ethylene oxide), Poly(ethyleneoxymethylene oxide),Poly(heptaldehyde), Poly(hexamethyleneoxide),Poly(hexamethyleneoxymethylene oxide), Poly(isobutyleneoxide),Poly(isobutyraldehyde), Poly(isophthalaldehyde),Poly(isopropylideneoxide), Poly(isovaleraldehyde),Poly(methyleneoxypentamethylene oxide),Poly(methyleneoxytetramethyleneoxide), Poly(methyleneoxynonamethyleneoxide), Poly(methyleneoxyoctamethylene oxide),Poly(methyleneoxytetradecamethyleneoxide), Poly(nonaldehyde),Poly(decamethylene oxide), Poly(nonamethyleneoxide),Poly(octamethyleneoxide), Poly(trimethylene oxide),Poly(3,3-bisazidomethyl-trimethyleneoxide),Poly(3,3-bischloromethyl-trimethyleneoxide),Poly(3,3-bisbromomethyl-trimethyleneoxide),Poly(3,3-bisethoxymethyl-trimethylene oxide),Poly(3,3-bisiodomethyl-trimethylene oxide),Poly(2,2-bistrifluoromethyl-trimethylene oxide),Poly(3,3-dimethyl-trimethylene oxide), Poly(3,3-diethyl-trimethyleneoxide), Poly(3-ethyl-3-methyl-trimethylene oxide), Poly(caprylaldehyde),Poly(propionaldehyde), Poly(3-methoxycarbonyl-propionaldehyde),Poly(3-cyano-propionaldehyde), Poly(propylene oxide),Poly(2-chloromethyl-propylene oxide), Poly[3-(1-naphthoxy)-propyleneoxide], Poly[3-(2-naphthoxy)-propylene oxide], Poly(3-phenoxy-propyleneoxide), Poly[3-(o-chloro-phenoxy)propylene oxide],Poly[3-(p-chloro-phenoxy)propylene oxide],Poly[3-(dimethyl-phenoxy)propylene oxide],Poly[3-(o-isopropyl-phenoxy)propylene oxide],Poly[3-(p-methoxy-phenoxy)propylene oxide],Poly[3-(m-methyl-phenoxy)propylene oxide],Poly[3-(o-methyl-phenoxy)propylene oxide],Poly[3-(o-phenyl-phenoxy)propylene oxide],Poly[3-(2,4,6-trichloro-phenoxy)propylene oxide],Poly(3,3,3-trifluoro-propylene oxide), Poly(tetramethylene oxide),Poly(cyclopropylidenedimethylene oxide), Poly(styrene oxide),Poly(allyloxymethylethylene oxide), Poly(butoxymethylethylene oxide),Poly(butylethyleneoxide), Poly(4-chlorobutylethyleneoxide),Poly(2-chloroethylethyleneoxide), Poly(2-cyanoethyloxymethyleneoxide),Poly(t-butylethylene oxide), Poly(2,2-bischloromethyltrimethyleneoxide), Poly(decylethylene oxide), Poly(ethoxymethylethyleneoxide),Poly(2-ethyl-2-chloromethyltrimethylene oxide),Poly(ethylethyleneoxide),Poly[1-(2,2,3,3,-tetrafluorocyclobutyl)ethylene oxide),Poly(octafluorotetramethylene oxide),Poly[1-(heptafluoro-2-propoxymethyl)ethylene], Poly(hexylethyleneoxide),Poly[(hexyloxymethyl)ethylene oxide],Poly(methyleneoxy-2,2,3,3,4,4-hexafluoro-pentamethylene oxide),Poly(methyleneoxy-2,2,3,3,4,4,5,5-octafluoro-hexamethylene oxide),Poly(1,1-dimethylethylene oxide), Poly(1,2-dimethylethylene oxide),Poly(1-methyltrimethylene oxide), Poly(2-methyltrimethylene oxide),Poly(methyleneoxytetramethylene oxide), Poly(octadecylethylene oxide),Poly(trifluoropropylene oxide),Poly(1,1-difluoroethyliminotetrafluoroethylene oxide),Poly(trifluoromethyliminotetrafluoro oxide), Poly(1,2-hexylene oxide),Poly(ethylenethioethylene oxide), Poly(difluoromethylene sulfide),Poly(methylenethiotetramethylene sulfide), Poly(1-ethylethylenesulfide), Poly(ethylmethylethylene sulfide),Poly(2-ethyl-2-methyltrimethylene sulfide),Poly(ethylene.trimethylene.sulfide), Poly(t-butylethylene sulfide),Poly(isopropylethylene sulfide), Poly(hexamethylene sulfide),Poly(1,2-cyclohexylene sulfide), Poly(1,3-cyclohexylene sulfide),Poly(1,2-cyclohexylene sulfone), Poly(1,3-cyclohexylene sulfone),Poly(hexamethylene sulfone), Poly(pentamethylene sulfide),Poly(pentamethylene sulfone), Poly(propylene sulfide), Poly(isobutylenesulfide), Poly(isopropylidene sulfide), Poly(2-butene sulfide),Poly(hexamethylenethiopentamethylene sulfide),Poly(hexamethylenethiotetramethylene sulfide), Poly(trimethylenesulfide), Poly(1-methyltrimethylene sulfide),Poly(3-methyl-6-oxo-hexamethylene sulfide),Poly(1-methyl-3-oxo-trimethylene sulfide), Poly(6-oxohexamethylenesulfide), Poly(2,2-dimethyl-trimethylene sulfide), Poly(trimethylenesulfone), Poly(2,2-dimethyltrimethylene sulfone),Poly(2,2-diethyltrimethylene sulfone), Poly(2,2-dipentyltrimethylenesulfone), Poly(tetramethylene sulfide), Poly(tetramethylene sulfone),Poly(ethylenethiohexamethylene sulfide), Poly(ethylenethiotetramethylenesulfide), Poly(pentamethylenethiotetramethylene sulfide),Poly(tetramethylene sulfide), Poly(decamethylene sulfide), Poly(p-tolylvinyl sulfoxide), Poly(decamethylene disulfide),Poly(heptamethylenedisulfide), Poly(hexamethylenedisulfide),Poly(nonamethylene disulfide), Poly(octamethylene disulfide),Poly(pentamethylene disulfide), Poly(octamethylenedithiotetramethylenedisulfide), Poly(oxyethylenedithioethylene),Poly(oxyethylenetetrathioethylene), Poly(dimethylketene),Poly(thiocarbonyl-3-methylpentamethylene),Poly(thiocarbonyl-2-methylpentamethylene),Poly(thiocarbonyl-1-methylethylene),Poly(thiocarbonyl-1-p-methoxybenzenesulfonylethylene),Poly(thiocarbonyl-1-tosylaminoethylene),Poly(thiocarbonyl-1-p-chlorobenzenesulfoamidoethylene),Poly(butylethyleneamine), Poly(ethylethylene amine),Poly(isobutylethylene amine), Poly(1,2-diethylethylene amine),Poly(1-butyl-2-ethylethylene amine), Poly(2-ethyl-1-pentylethylene),Poly(N-formyl-isopropylethylene), Poly(isopropylethylene amine),Poly(N-formylpropylene amine), Poly(ethylene trimethylene amine),Poly(N-acetyl-ethylene amine), Poly(N-benzoyl-ethylene amine),Poly[N-(p-chloro-benzoyl)-ethylene amine],Poly(N-butyryl-ethyleneamine),Poly[N-[4-(4-methylthiophenoxy)-butyryl]-ethyleneamine],Poly(N-cyclohexanecarbonyl-ethylene amine), Poly(N-dodecanoyl-ethyleneamine), Poly(N-heptanoyl-ethyleneamine), Poly(N-hexanoyl-ethyleneamine),Poly(N-isobutyryl-ethylene amine), Poly(N-isovaleryl-ethylene amine),Poly(N-octanoyl-ethylene amine), Poly(N-2-naphthoyl-ethylene amine),Poly(N-p-toluoyl-ethylene amine), Poly(N-perfluorooctaoyl-ethyleneamine), Poly(N-perfluoropropionyl-ethylene amine),Poly(N-pivaloyl-ethyleneamine), Poly(N-valeryl-ethyleneamine),Poly(trimethyleneamine), Polysilane, Poly(di-N-hexyl-silane),Poly(di-N-pentyl-silane), Poly(vinyltriethoxysilane),Poly(vinyltrimethoxysilane), Poly(vinyltrimethylsilane), Poly(vinylmethyldiacetoxysilane), Poly(vinyl methyldiethoxysilane),Poly(vinylphenyldimethylsilane), Polysiloxane, Poly(diethylsiloxane),Poly(dimethylsiloxane), Poly(diphenylsiloxane), Poly(dipropylsiloxane),Poly(pentaphenyl-p-toluoyltrsiloxane), Poly(phenyl-p-toluoylsiloxane),Polytphthalocyaninato-siloxane), Poly(propylmethylsiloxane),Poly(ethylmethylsiloxane), Poly(methyloctylsiloxane),Poly(3,3,3-trifluoropropylmethylsiloxane), Poly(vinylmethylsiloxane),Polysilylene, Poly(dimethylsilylene), Poly(diphenylsilylene),Poly(dimethyldiallylsilane),Poly[oxydi(pentafluorophenyl)silylenedi(oxydimethylsilylene)],Poly[oxymethylchlorotetrafluorophenylsilylenedi(oxydimethylsilylene)],Poly(oxymethylpentafluorophenylsilylene),Poly(oxymethylpentafluorophenylsilyleneoxydimethylsilylene,Poly[oxymethylpentafluorophenylsilylenedi(oxydimethylsilylene)],Poly(oxymethyl-3,3,3-trifluoropropylsilylene),Poly(oxymethylphenylsilylene),Poly[tri(oxydimethylsilylene)oxy(methyl)trimethylsiloxysilylene],Poly[tri(oxydimethylsilylene)oxy(methyl)-2-phenyl-ethylsilylene],Poly[(4-dimethylaminophenyl)methylsilylenetrimethylene],Poly[(4-dimethylaminophenyl)phenylsilylenetrimethylene],Poly[(methyl)phenylsilylenetrimethylene], Poly(1,1-dimethylsilazane),Poly(dimethylsilylenetrimethylene),Poly(di-p-tolylsilylenetrimethylene), Poly(phosphazene),Poly(bis-beta-naphthoxy-phosphazene), Poly(bis-phenoxy-phosphazene),Poly(di-p-methyl-bis-phenoxy-phosphazene),Poly(di-p-chloro-bis-phenoxy-phosphazene),Poly(di-2,4-dichloro-bis-phenoxy-phosphazene),Poly(di-p-phenyl-bis-phenoxy-phosphazene),Poly(di-m-trifluoromethyl-phosphazene), Poly(di-methyl-phosphazene),Poly(dich-oro-phosphazene), Poly(diethoxy-phosphazene),Poly[bis(ethylamino)phosphazene],Poly[bis(2,2,2-trifluoroethoxy)phosphazene],Poly[bis(3-trifluoromethylphenoxy)phosphazene],Poly[bis(1H,1H-pentadecafluorooctyloxy)phosphazene],Poly[bis(1H,1H-pentafluoropropoxy)phosphazene],Poly(dimethoxy-phosphazene), Poly[bis(phenylamino)phosphazene],Poly[bis(piperidino)phosphazene], Poly(diethylpropenyl phosphate),Poly(diethylisopropenyl phosphate), Poly[vinylbis(chloroethyl)phosphate], Poly(vinyldisethylphosphate),Poly(vinyldiethyl phosphate), Poly(vinyldiphenyl phosphate),Poly(alpha-bromovinyl diethyl phosphonate), Poly(alpha-carboethoxyvinyldiethyl phosphonate), Poly(alpha-carbomethoxyvinyl diethyl phosphonate),Poly(isopropenyl dimethyl phosphonate), Poly[vinylbis(2-chloroethyl)phosphonate], Poly(vinyl dibutyl phosphonate),Poly(vinyl diethyl phosphonate), Poly(vinyldiisobutyl phosphonate),Poly(vinyl diisopropyl phosphonate), Poly(vinyl dimethyl phosphonate),Poly(vinyl diphenyl phosphonate), Poly(vinyl dipropyl phosphonate),Poly[2-(4-vinylphenyl)ethyl diethyl phosphonate), Poly(4-vinylphenyldiethyl phosphonate), and Poly(diphenylvinyl phosphine oxide).

Method of Manufacture

A method of responsive microgel synthesis and production is further anobject of the present invention. The method of the present inventioninvolves a single synthetic step, which is advantageous for scale-up ofresponsive microgel fabrication. The synthesis of the microgelsdescribed herein involves a free-radical copolymerization of a vinylmonomer with a divinyl cross-linker with simultaneous hydrogenabstraction from a polymer present in the reaction system. The hydrogenabstraction leads to generation of macro-radicals that lead to thegrafting of the amphiphilic copolymer ‘dangling chains’ onto the growingmicrogel network. The series of reactions that occur simultaneously andyield a responsive microgel of the present invention are shown in FIG. 4(scheme of the one-step synthesis of responsive microgels). See, e.g.,Examples I and II.

A preferred chain-transfer reaction to covalently bond the nonioniccopolymer to the ionizable network is a free-radical polymerization(using a redox free-radical initiator) of an ionizable monomer and adivinyl cross-linker.

A method of making the responsive microgel covalently cross-linkedpolymer network (graft-comb copolymer) of the present invention, forexample, comprises the steps of: a) providing, an ionizable monomer, adivinyl cross-linker, a free radical, and a nonionic copolymer; and, b)copolymerizing the ionizable monomer with the divinyl cross-linker toproduce an ionizable network, while c) abstracting hydrogen from thenonionic copolymer with the free radical to progress a chain transferreaction wherein the nonionic copolymer is covalently bonded onto theionizable network to produce a responsive microgel as defined herein.

Divinyl cross-linker as used herein refers to a reactive chemical havingat least two ethylenic double bonds capable of participating in at leasttwo growing polymer chains. Examples of cross-linkers of this type,which are normally used as crosslinkers in polymerization reactions, areN,N′-methylenebisacrylamide, polyethylene glycol diacrylates andpolyethylene glycol dimethacrylates which are derived in each case frompolyethylene glycols with a molecular weight of from 106 to 8500,preferably 400 to 2000, trimethylolpropane triacrylate,trimethylolpropane trimethacrylate, ethylene glycol diacrylate,propylene glycol diacrylate, butanediol diacrylate, hexanedioldiacrylate, hexanediol dimethacrylate, diacrylates and dimethacrylatesof block copolymers of ethylene oxide and propylene oxide, polyhydricalcohols such as glycerol or pentaerythritol which are esterified two orthree times with acrylic acid or methacrylic acid, triallylamine,tetraallylethylenediamine, divinylbenzene, diallyl phthalate,polyethylene glycol divinyl ethers of polyethylene glycols with amolecular weight of from 126 to 4000, trimethylolpropane diallyl ether,butanediol divinyl ether, pentaerythritol triallyl ether and/ordivinylethyleneurea. Water-soluble crosslinkers are preferably used,e.g. N,N′-methylenebisacrylamide, oligoethylene glycol diacrylates andoligoethylene glycol dimethacrylates derived from adducts of 2 to 400mol of ethylene oxide and 1 mol of a diol or polyol, vinyl ethers ofadducts of 2 to 400 mol of ethylene oxide and 1 mol of a diol or polyol,ethylene glycol diacrylate, ethylene glycol dimethacrylate ortriacrylates and trimethacrylates of adducts of 6 to 20 mol of ethyleneoxide and one mol of glycerol, pentaerythritol triallyl ether and/ordivinylurea.

Also suitable as crosslinkers are compounds, which contain at least onepolymerizable ethylenically unsaturated group and at least one otherfunctional group. The functional group in these crosslinkers must beable to react with the functional groups, essentially the carboxylgroups in the monomers of the backbone. Examples of suitable functionalgroups are hydroxyl, amino, epoxy and aziridino groups.

Also suitable as crosslinkers are those compounds which have at leasttwo functional groups able to react with carboxyl and other functionalgroups in the monomers used. The suitable functional groups have alreadybeen mentioned above, i.e. hydroxyl, amino, epoxy, isocyanate, ester,amide and aziridino groups. Examples of such crosslinkers are ethyleneglycol, diethylene glycol, triethylene glycol, tetraethylene glycol,polyethylene glycol, glycerol, polyglycerol, propylene glycol,polypropylene glycol, block copolymers of ethylene oxide and propyleneoxide, sorbitan fatty acid esters, ethoxylated sorbitan fatty acidesters, trimethylolpropane, pentaerythritol, polyvinyl alcohol,sorbitol, polyglycidyl ethers such as ethylene glycol diglycidyl ether,polyethylene glycol diglycidyl ether, glycerol diglycidyl ether,glycerol polyglycidyl ether, diglycerol polyglycidyl ether, polyglycerolpolyglycidyl ether, sorbitol polyglycidyl ether, pentaerythritolpolyglycidyl ether, propylene glycol diglycidyl ether and polypropyleneglycol diglycidyl ether, polyaziridine compounds such as2,2-bishydroxymethylbutanol tris[3-(1-aziridinyl)propionate],1,6-hexamethylenediethyleneurea,4,4′-methylenebis(phenyl)-N,N′-diethyleneurea, halo epoxy compounds suchas epichlorohydrin and a-methylfluorohydrin, polyisocyanates such as2,4-tolylene diisocyanate and hexamethylene diisocyanate, alkylenecarbonates such as 1,3-di-oxolan-2-one and 4-methyl-1,3-dioxolan-2-one,polyquaternary amines such as condensates of dimethylamine withepichlorohydrin, homo- and copolymers of diallyldimethylammoniumchloride, and homo- and copolymers of dimethylaminoethyl (meth)acrylate,which are, where appropriate, quaternized with, for example, methylchloride.

Other suitable crosslinkers are polyvalent metal ions able to form ioniccrosslinks. Examples of such crosslinkers are magnesium, calcium, bariumand aluminum ions. A preferred crosslinker of this type is sodiumaluminate. These crosslinkers are added, for example, as hydroxides,carbonates or bicarbonates to the aqueous polymerizable solution.

Other suitable crosslinkers are multifunctional bases which are likewiseable to form ionic crosslinks, for example polyamines or theirquaternized salts. Examples of polyamines are ethylenediamine,diethylenetriamine, triethylenetetramine, tetraethylenepentamine,pentaethylenehexamine and polyethyleneimines, and polyvinylamines withmolecular weights of up to 4,000,000 in each case.

In a preferred embodiment of the invention, divinyl crosslinkers areused. These can be hydrophobic or most preferably amphiphilic orhydrophilic. Apart from polyvalent metal ions, all the water-insolublecrosslinkers which are described above and can be assigned to thevarious groups are suitable for producing gels. Some preferredhydrophobic crosslinkers are diacrylates or dimethacrylates or divinylethers of alkanediols with 2 to 25 carbon atoms (branched, linear, withany suitable arrangement of OH groups) such as 1,3-propanediol,1,4-butanediol, 1,6-hexanediol, neopentyl glycol, 1,9-nonanediol or1,2-dodecanediol, di-, tri- or polypropylene glycol diacrylates ordimethacrylates, allyl acrylate, allyl methacrylate, divinylbenzene,glycidyl acrylate or glycidyl methacrylate, allyl glycidyl ether andbisglycidyl ethers of the alkanediols listed above.

Examples of suitable hydrophilic crosslinkers areN,N′-methylenebisacrylamide, polyethylene glycol diacrylates ordimethacrylates with a molecular weight from 200 to 4000, divinylurea,triallylamine, diacrylates or dimethacrylates of adducts of from 2 to400 mol of ethylene oxide and 1 mol of a diol or polyol or thetriacrylate of an adduct of 20 mol of ethylene oxide and 1 mol ofglycerol and vinyl ethers of adducts of from 2 to 400 mol of ethyleneoxide and 1 mol of a diol or polyol.

The polymerization initiators which can be used are all initiators whichform free radicals under the polymerization conditions and which arenormally used in the preparation of responsive gels. It is also possibleto initiate the polymerization by the action of electron beams on thepolymerizable aqueous mixture. However, the polymerization can also bestarted in the absence of initiators of the abovementioned type by theaction of high-energy radiation in the presence of photoinitiators.

Polymerization initiators which can be used are all compounds whichdecompose to free radicals under the polymerization conditions, eg.peroxides, hydroperoxides, hydrogen peroxide, persulfates, azo compoundsand the redox catalysts. Initiators soluble in the mixture of themonomer and amphiphilic copolymer are preferably used. It isadvantageous in some cases to use mixtures of various polymerizationinitiators, e.g. most preferably mixtures of lauroyl peroxide or benzoylperoxide hydrogen peroxide with 2,2′-azobis(2,4-dimethylpentanenitrile)or 4,4′-azobis(4-cyanovaleric acid). Examples of suitable organicperoxides are acetylacetone peroxide, methyl ethyl ketone peroxide,tertbutyl hydroperoxide, cumene hydroperoxide, tert-amyl perpivalate,tert-butyl perpivalate, tert-butyl perneohexanoate, tert-butylperisobutyrate, tert-butyl per-2-ethylhexanoate, tert-butylperisononanoate, tert-butyl permaleate, tert-butyl perbenzoate,di(2-ethylhexyl)peroxydicarbonate, dicyclohexyl peroxydicarbonate,di(4-tert-butylcyclohexyl)peroxydicarbonate, dimyristylperoxydicarbonate, diacetyl peroxydicarbonate, allyl peresters, cumylperoxyneodecanoate, tert-butyl per-3,5,5-trimethylhexanoate, acetylcyclohexylsulfonyl peroxide, dilauroyl peroxide, dibenzoyl peroxide andtert-amyl perneodecanoate. Also suitable polymerization initiators arewater-soluble azo initiators, eg.2,2′-azobis(2-amidinopropane)dihydrochloride,2,2′-azobis(N,N′-dimethyleneisobutyramidine)dihydrochloride,2-(carbamoylazo)isobutyronitrile,2,2′-azobis[2-(2-imidazolin-2-yl)propane]dihydrochloride and4,4,-azobis(4-cyanovaleric acid). Polymerization initiators are used inconventional amounts, e.g. in amounts of from 0.01 to 5, preferably 0.1to 2.0, % of the weight of the monomers to be polymerized.

Also suitable as initiators are redox catalysts. The redox catalystscontain as oxidizing component at least one of the abovementioned peroxycompounds and as reducing component, for example, ascorbic acid,glucose, sorbose, ammonium or alkali metal bisulfite, sulfite,thiosulfate, hyposulfite, pyrosulfite or sulfide, metal salts such asiron(II) ions or silver ions, or sodium hydroxymethylsulfoxylate. Thereducing component preferably used in the redox catalyst is ascorbicacid or sodium sulfite. Based on the amount of monomers used in thepolymerization, for example, from 3×10⁻⁶ to 1 mol % of the reducingcomponent of the redox catalyst system and from 0.001 to 5.0 mol % ofthe oxidizing component of the redox catalyst are used.

If the polymerization is initiated by the action of high-energyradiation, photoinitiators are normally used as initiator. These may be,for example, alpha-splitters, H-abstracting systems or else azides.Examples of initiators of these types are benzophenone derivatives suchas Michler's ketone, phenanthrene derivatives, fluorene derivatives,anthraquinone derivatives, thioxanthone derivatives, coumarinderivatives, benzoin ethers and derivatives thereof, azo compounds likethe free-radical formers mentioned above, substitutedhexaarylbisimidazoles or acylphosphine oxides. Examples of azides are:2-(N,N-dimethylamino)ethyl 4-azidocinnamate,2-(N,N-dimethylamino)ethyl-4-azidonaphthyl ketone,2-(N,N-dimethylamino)ethyl 4-azidobenzoate, 5-azido-1-naphthyl2-(N,N-dimethylamino)ethyl sulfone, N-(4-sulfonylazidophenyl)maleimide,N-acetyl-4-sulfonylazidoaniline, 4-sulfonylazido-aniline,4-azidoaniline, 4-azidophenacyl bromide, p-azidobenzoic acid,2,6-bis(p-azidobenzylidene)cyclohexanone and2,6-bis(p-azidobenzylidene)-4-methylcyclohexanone. The photoinitiatorsare, if employed, normally used in amounts of from 0.01 to 5% of theweight of the monomers to be polymerized.

In one embodiment, it is preferred to use free-radical initiatorscapable of abstracting tertiary and secondary hydrogens from thebackbone of the amphiphilic polymer of the present invention.

Methods of Use

One of the major concerns in the delivery of drugs is thebioavailability of the drug. Depending upon the nature of the drug andthe route of delivery, the bioavailability may be very low due to, forexample, the degradation of oral-delivered drugs byhepato-gastrointestinal first-pass elimination or rapid clearance of thedrug from the site of application. The net result is that frequentdosing may be required with higher than needed amounts of drug, whichcan lead to undesired side effects. Thus, it is desired by thepharmaceutical industry to have ways of administering drugs such thattheir availability can be controlled in an even dosing manner, theamounts of drugs can be kept as low as possible to minimize sideeffects, and dosing regime can be kept to a minimum to provide greaterconvenience to the subject, thus promoting greater compliance withappropriate dosing.

The responsive microgels of the present invention are useful in a widevariety of chemo-mechanical applications in that they display diversephase transition characteristics. A method, for example, of deliveringat least one therapeutic or cosmetic agent to a mammalian subject is apreferred embodiment of the invention which comprises administering aresponsive microgel of the present invention to the subject whichcomprises at least one such agent.

A method of delivering an effective amount at least one therapeuticagent to a patient is a preferred method of the invention whichcomprises administering an effective amount of a responsive microgel ofthe present invention which comprises at least one therapeutic agent.Therapeutic regimens for the prevention and/or treatment of cancerfrequently requires, for example, the administration of an effectiveamount of a cationic, hydrophobic, and/or amphiphilic compound,individually or in combinations. The responsive microgel of the presentinvention is particularly suited for therapeutic administration of thesetypes of agents or entities. The responsive microgels are provided as along-term delivery device for therapeutic agents and to enhance thetherapeutic profile. The responsive microgels provide improved andsubstantially linear sustained release of therapeutic agents to improveand prolong the bioavailability of the agent. The reversibly gellingresponsive microgel of this invention has the physico-chemicalcharacteristics that make it a suitable delivery vehicle forconventional small chemical drugs as well as new macromolecular (e.g.,peptides) drugs or therapeutic products.

The responsive microgel of the present invention is particularly suitedfor oral administration. The responsive microgel of the presentinvention may also be employed to deliver therapeutic entities(including cosmetic agents), for example, by intranasal, ocular,pulmonary, colonic, vaginal, as well as topical administration. Thetemperature-responsive mode of solute solubilization, for example, bymicrogels of the present invention is useful for medicinal as well ascosmetic formulations. Preferred therapeutic entities for use in thepresent invention include but are not limited to doxorubicin,mitoxantrone, mitomycin C, as well as the Taxanes including but notlimited to (paclitaxel (TAXOL®), and docetaxel (TAXOTERE®)).

Examples of therapeutic entities that might be utilized in a deliveryapplication of the invention include literally any hydrophilic orhydrophobic biologically active compound. Preferably, though notnecessarily, the drug is one that has already been deemed safe andeffective for use by the appropriate governmental agency or body. Forexample, drugs for human use listed by the FDA under 21 C.F.R. 330.5,331 through 361; 440-460; drugs for veterinary use listed by the FDAunder 21 C.F.R. 500-582, incorporated herein by reference, are allconsidered acceptable for use in the present responsive microgel.

Drugs that are not themselves liquid at body temperature can beincorporated into the responsive microgel of the present invention.Moreover, peptides and proteins which may normally be rapidly degradedby tissue-activated enzymes such as peptidases, can be passivelyprotected in the microgels described herein.

A responsive microgel which comprises at least one therapeutic entity isparticularly preferred. A responsive microgel which comprises at leastone anticancer agent is a preferred embodiment of the present inventionwherein, for example, at least one anticancer agent is selected from thegroup consisting of (a steroidal antiandrogen, a non steroidalantiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, aselective estrogen receptor modulator (SERM), a taxane, and a LHRHanalog). Non steroidal antiandrogen as referred to herein includes butis not limited to the group consisting essentially of (finasteride(PROSCAR®), flutamide (4′-nitro-3′-trifluorormethyl isobutyranilide),bicalutamide (CASODEX®), and nilutamide). SERM as referred to hereinincludes but is not limited to the group consisting essentially of(tamoxifen, raloxifene, droloxifene, and idoxifene). LHRH analog asreferred to herein includes but is not limited to the group consistingessentially of (goserelin acetate (ZOLADEX®), and leuprolide acetate(LUPRON®)).

A method of prevention or treatment of a tumor is provided comprisingadministering a therapeutically effective amount of a responsivemicrogel which comprises at least one therapeutic entity to a patientwherein the patient is either at risk of developing a tumor or alreadyexhibits a tumor. A method of prevention or treatment of a tumor isprovided wherein at least one agent described herein—or a stereoisomericmixture thereof, diastereomerically enriched, diastereomerically pure,enantiomerically enriched or enantiomerically pure isomer thereof, or aprodrug of such compound, mixture or isomer thereof, or apharmaceutically acceptable salt of the compound, mixture, isomer orprodrug—is administered in a therapeutically effective amount comprisedwithin a responsive microgel of the present invention to a patientwherein the patient is either at risk of developing a tumor or alreadyexhibits a tumor. Methods of employing the responsive microgel of thepresent invention for the prevention or treatment of a tumor is providedwherein at least one agent is comprised within the microgel selectedfrom the group consisting of (a steroidal antiandrogen, a non steroidalantiandrogen, an estrogen, diethylstilbestrol, a conjugated estrogen, aselective estrogen receptor modulator (SERM), a taxane, and a LHRHanalog) and an effective amount of the microgel is administered to apatient in need of treatment.

The term therapeutic entity includes pharmacologically active substancesthat produce a local or systemic effect in a mammal. The term thus meansany substance intended for use in the diagnosis, cure, mitigation,treatment or prevention of disease or in the enhancement of desirablephysical or mental development and conditions in a mammal.

Therapeutic entities for employment with the responsive microgelsdescribed herein therefore include small molecule compounds,polypeptides, proteins, nucleic acids, and PLURONIC®, for example, asdescribed herein (e.g., and for the formation of mixed micelles).

Examples of proteins include antibodies, enzymes, growth hormone andgrowth hormone-releasing hormone, gonadotropin-releasing hormone, andits agonist and antagonist analogues, somatostatin and its analogues,gonadotropins such as luteinizing hormone and follicle-stimulatinghormone, peptide-T, thyrocalcitonin, parathyroid hormone, glucagon,vasopressin, oxytocin, angiotensin I and II, bradykinin, kallidin,adrenocorticotropic hormone, thyroid stimulating hormone, insulin,glucagon and the numerous analogues and congeners of the foregoingmolecules.

Classes of pharmaceutically active compounds which can be loaded ontoresponsive microgel compositions of the invention include, but are notlimited to, anti-AIDS substances, anti-cancer substances, antibiotics,immunosuppressants (e.g. cyclosporine) anti-viral substances, enzymeinhibitors, neurotoxins, opioids, hypnotics, antihistamines,tranquilizers, anti-convulsants, muscle relaxants and anti-Parkinsonsubstances, anti-spasmodics and muscle contractants, miotics andanti-cholinergics, antiglaucoma compounds, anti-parasite and/oranti-protozoal compounds, anti-hypertensives, analgesics, anti-pyreticsand anti-inflammatory agents such as NSAIDs, local anesthetics,ophthalmics, prostaglandins, anti-depressants, anti-psychoticsubstances, anti-emetics, imaging agents, specific targeting agents,neurotransmitters, proteins, cell response modifiers, and vaccines.

A more complete listing of classes of compounds suitable for loadinginto polymers using the present methods may be found in thePharmazeutische Wirkstoffe (Von Kleemann et al. (eds) Stuttgart/NewYork, 1987, incorporated herein by reference). Examples of particularpharmaceutically active substances are presented below:

Anti-AIDS substances are substances used to treat or prevent AutoimmuneDeficiency Syndrome (AIDS). Examples of such substances include CD4,3′-azido-3′-deoxythymidine (AZT), 9-(2-hydroxyethoxymethyl)-guanineacyclovir( ), phosphonoformic acid, 1-adamantanamine, peptide T, and2′,3′ dideoxycytidine.

Anti-cancer substances are substances used to treat or prevent cancer.Examples of such substances include methotrexate, cisplatin, prednisone,hydroxyprogesterone, medroxyprogesterone acetate, megestrol acetate,diethylstilbestrol, testosterone propionate, fluoxymesterone,vinblastine, vincristine, vindesine, daunorubicin, doxorubicin,hydroxyurea, procarbazine, aminoglutethimide, mechlorethamine,cyclophosphamide, melphalan, uracil mustard, chlorambucil, busulfan,carmustine, lomusline, dacarbazine (DTIC:dimethyltriazenomidazolecarboxamide), methotrexate, fluorouracil,5-fluorouracil, cytarabine, cytosine arabinoxide, mercaptopurine,6-mercaptopurine, thioguanine.

Antibiotics are art recognized and are substances which inhibit thegrowth of or kill microorganisms. Antibiotics can be producedsynthetically or by microorganisms. Examples of antibiotics includepenicillin, tetracycline, chloramphenicol, minocycline, doxycycline,vanomycin, bacitracin, kanamycin, neomycin, gentamycin, erythromicin andcephalosporins.

Anti-viral agents are substances capable of destroying or suppressingthe replication of viruses. Examples of anti-viral agents includea-methyl-P-adamantane methylamine, 1,-D-ribofuranosyl-1,2,4-triazole-3carboxamide, 9->2-hydroxy-ethoxy!methylguanine, adamantanamine,5-iodo-2′-deoxyuridine, trifluorothymidine, interferon, and adeninearabinoside.

Enzyme inhibitors are substances which inhibit an enzymatic reaction.Examples of enzyme inhibitors include edrophonium chloride,N-methylphysostigmine, neostigmine bromide, physostigmine sulfate,tacrine HCl, tacrine, 1-hydroxy maleate, iodotubercidin,p-bromotetramisole, 10-(alpha-diethylaminopropionyl)-phenothiazinehydrochloride, calmidazolium chloride,hemicholinium-3,3,5-initrocatechol, diacylglycerol kinase inhibitor I,diacylglycerol kinase inhibitor II, 3-phenylpropargylamine,N.sup.6-monomethyl-L-arginine acetate, carbidopa,3-hydroxybenzylhydrazine HCl, hydralazine HCl, clorgyline HCl, deprenylHCl,L(−)-, deprenyl HCl,D(+)-, hydroxylamine HCl, iproniazid phosphate,6-MeO-tetrahydro-9H-pyrido-indole, nialamide, pargyline HCl, quinacrineHCl, semicarbazide HCl, tranylcypromine HCl,N,N-diethylaminoethyl-2,2-diphenylvalerate hydrochloride,3-isobutyl-1-methylxanthne, papaverine HCl, indomethacind,2-cyclooctyl-2-hydroxyethylamine hydrochloride,2,3-dichloro-a-methylbenzylamine PCMB),8,9-dichloro-2,3,4,5-tetrahydro-1H-2-benzazepine hydrochloride,p-aminoglutethimide, p-aminoglutethimide tartrate,R(+)-,p-aminoglutethimide tartrate,S(−)-, 3-iodotyrosine,alpha-methyltyrosine,L-, alpha-methyltyrosine,D L-, acetazolamide,dichlorphenamide, 6-hydroxy-2-benzothiazolesulfonamide, and allopurinol.

Neurotoxins are substances which have a toxic effect on the nervoussystem, e.g. nerve cells. Neurotoxins include adrenergic neurotoxins,cholinergic neurotoxins, dopaminergic neurotoxins, and otherneurotoxins. Examples of adrenergic neurotoxins includeN-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride. Examples ofcholinergic neurotoxins include acetylethylcholine mustardhydrochloride. Examples of dopaminergic neurotoxins include6-hydroxydopamine HBr,1-methyl-4-(2-methylphenyl)-1,2,3,6-tetrahydro-pyridine hydrochloride,1-methyl-4-phenyl-2,3-dihydropyridinium perchlorate,N-methyl-4-phenyl-1,2,5,6-tetrahydropyridine HCl,1-methyl-4-phenylpyridinium iodide.

Opioids are substances having opiate like effects that are not derivedfrom opium. Opioids include opioid agonists and opioid antagonists.Opioid agonists include codeine sulfate, fentanyl citrate, hydrocodonebitartrate, loperamide HCl, morphine sulfate, noscapine, norcodeine,normorphine, thebaine. Opioid antagonists include nor-binaltorphimineHCl, buprenorphine, chlornaltrexamine 2HCl, funaltrexamione HCl,nalbuphine HCl, nalorphine HCl, naloxone HCl, naloxonazine, naltrexoneHCl, and naltrindole HCl.

Hypnotics are substances which produce a hypnotic effect. Hypnoticsinclude pentobarbital sodium, phenobarbital, secobarbital, thiopentaland mixtures, thereof, heterocyclic hypnotics, dioxopiperidines,glutarimides, diethyl isovaleramide, a-bromoisovaleryl urea, urethanesand disulfanes.

Antihistamines are substances which competitively inhibit the effects ofhistamines. Examples include pyrilamine, chlorpheniramine,tetrahydrazoline, and the like.

Lubricants are substances that increase the lubricity of the environmentinto which they are delivered. Examples of biologically activelubricants include water and saline.

Tranquilizers are substances which provide a tranquilizing effect.Examples of tranquilizers include chloropromazine, promazine,fluphenzaine, reserpine, deserpidine, and meprobamate.

Anti-convulsants are substances which have an effect of preventing,reducing, or eliminating convulsions. Examples of such agents includeprimidone, phenyloin, valproate, Chk and ethosuximide.

Muscle relaxants and anti-Parkinson agents are agents which relaxmuscles or reduce or eliminate symptoms associated with Parkinson'sdisease. Examples of such agents include mephenesin, methocarbomal,cyclobenzaprine hydrochloride, trihexylphenidyl hydrochloride,levodopa/carbidopa, and biperiden.

Anti-spasmodics and muscle contractants are substances capable ofpreventing or relieving muscle spasms or contractions. Examples of suchagents include atropine, scopolamine, oxyphenonium, and papaverine.

Miotics and anti-cholinergics are compounds which cause bronchodilation.Examples include echothiophate, pilocarpine, physostigmine salicylate,diisopropylfluorophosphate, epinephrine, neostigmine, carbachol,methacholine, bethanechol, and the like.

Anti-glaucoma compounds include betaxalol, pilocarpine, timolol, timololsalts, and combinations of timolol, and/or its salts, with pilocarpine.

Anti-parasitic, -protozoal and -fungals include ivermectin,pyrimethamine, trisulfapyrimidine, clindamycin, amphotericin B,nystatin, flucytosine, natamycin, and miconazole.

Anti-hypertensives are substances capable of counteracting high bloodpressure. Examples of such substances include alpha-methyldopa and thepivaloyloxyethyl ester of alpha-methyldopa.

Analgesics are substances capable of preventing, reducing, or relievingpain. Examples of analgesics include morphine sulfate, codeine sulfate,meperidine, and nalorphine.

Anti-pyretics are substances capable of relieving or reducing fever andanti-inflammatory agents are substances capable of counteracting orsuppressing inflammation. Examples of such agents include aspirin(salicylic acid), indomethacin, sodium indomethacin trihydrate,salicylamide, naproxen, colchicine, fenoprofen, sulindac, diflunisal,diclofenac, indoprofen and sodium salicylamide.

Local anesthetics are substances which have an anesthetic effect in alocalized region. Examples of such anesthetics include procaine,lidocain, tetracaine and dibucaine.

Ophthalmics include diagnostic agents such as sodium fluorescein, rosebengal, methacholine, adrenaline, cocaine, and atropine. Ophthalmicsurgical additives include alpha-chymotrypsin and hyaluronidase.

Prostaglandins are art recognized and are a class of naturally occurringchemically related, long-chain hydroxy fatty acids that have a varietyof biological effects.

Anti-depressants are substances capable of preventing or relievingdepression. Examples of anti-depressants include imipramine,amitriptyline, nortriptyline, protriptyline, desipramine, amoxapine,doxepin, maprotiline, tranylcypromine, pheneizine, and isocarboxazide.

Anti-psychotic substances are substances which modify psychoticbehavior. Examples of such agents include phenothiazines, butyrophenonesand thioxanthenes.

Anti-emetics are substances which prevent or alleviate nausea orvomiting. An example of such a substance includes dramamine.

In topical skin care applications, a variety of active substances may beadvantageously employed. By way of example only suitable active agentswhich may be incorporated into the cosmetic composition includeanti-aging active substances, anti-wrinkle active substances, hydratingor moisturizing or slimming active substances, depigmenting activesubstances, substances active against free radicals, anti-irritationactive substances, sun protective active substances, anti-acne activesubstances, firming-up active substances, exfoliating active substances,emollient active substances, and active substances for the treating ofskin disorders such as dermatitis and the like.

Imaging agents are agents capable of imaging a desired site, e.g. tumor,in vivo. Examples of imaging agents include substances having a labelwhich is detectable in vivo, e.g. antibodies attached to fluorescentlabels. The term antibody includes whole antibodies or fragmentsthereof.

Specific targeting agents include agents capable of delivering atherapeutic agent to a desired site, e.g. tumor, and providing atherapeutic effect. Examples of targeting agents include agents whichcan carry toxins or other agents which provide beneficial effects. Thetargeting agent can be an antibody linked to a toxin, e.g. ricin A or anantibody linked to a drug.

Neurotransmitters are substances which are released from a neuron onexcitation and travel to either inhibit or excite a target cell.Examples of neurotransmitters include dopamine, serotonin,q-aminobutyric acid, norepinephrine, histamine, acetylcholine, andepinephrine.

Cell response modifiers are chemotactic factors such as platelet-derivedgrowth factor (PDGF). Other chemotactic factors includeneutrophil-activating protein, monocyte chemoattractant protein,macrophage-inflammatory protein, platelet factor, platelet basicprotein, and melanoma growth stimulating activity; epidermal growthfactor, transforming growth factor (alpha), fibroblast growth factor,platelet-derived endothelial cell growth factor, insulin-like growthfactor, nerve growth factor, and bone growth/cartilage-inducing factor(alpha and beta), or other bone morphogenetic protein.

Other cell response modifiers are the interleukins, interleukininhibitors or interleukin receptors, including interleukin 1 throughinterleukin 10; interferons, including alpha, beta and gamma;hematopoietic factors, including erythropoietin, granulocyte colonystimulating factor, macrophage colony stimulating factor andgranulocyte-macrophage colony stimulating factor; tumor necrosisfactors, including alpha and beta; transforming growth factors (beta),including beta-1, beta-2, beta-3, inhibin, and activin; and bonemorphogenetic proteins.

As those skilled in the art will appreciate, the foregoing list isexemplary only. Because the responsive microgel of the present inventionis suited for application under a variety of physiological conditions, awide variety of pharmaceutical agents may be loaded onto the responsivemicrogels described herein and administered.

Formulations

Tablet Excipients. It has been demonstrated that standard pharmaceuticalprocesses, such as lyophilization and air-drying can process theresponsive microgel of the invention. The reversible thermalviscosifying responsive microgel may be reconstituted with water,phosphate buffer or calcium chloride solution, without loss ordegradation of rheological properties. Thus, it is contemplated that theresponsive microgel of the invention may also be incorporated asexcipients into tablets or granules for oral delivery, for example. Theresponsive microgel may be coated on an outer surface of the tablet ormay be introduced in powder form into the tablet along with the activeagent and other ingredients. The poloxamer:poly(acrylic acid)composition may be used to promote bioadhesion of the tablet and itscontents with the mucosal lining of the gastro-intestinal tract toextend transit time.

Also, when incorporated as a powder, the slow dissolution rate of theend-modified responsive microgel makes it a suitable excipient tosustained release tableting formulation. The addition of such responsivemicrogel would yield to a slow release of the incorporated drug.

Injectibles. The end-modified responsive microgel composition of theinvention is well-suited for use in injectable applications. A depotformulation may be prepared and administered at low viscosity to asubdermal or intramuscular site, for example. The responsive microgelwill viscosify and form a depot site, which will slowly release theactive agent. The reversible thermally viscosifying responsive microgel,upon contact with body fluids including blood or the like, undergoesgradual release of the dispersed drug for a sustained or extended period(as compared to the release from an isotonic saline solution). This canresult in prolonged delivery (over, say 1 to 2,000 hours, preferably 2to 800 hours) of effective amounts (say, 0.0001 mg/kg/hour to 10mg/kg/hour) of the drug. This dosage form can be administered as isnecessary depending on the subject being treated, the severity of theaffliction, the judgment of the prescribing physician, and the like.

Preparation of Pharmaceutic Compositions May be Accomplished withReference to any of the pharmaceutic formulation guidebooks and industryjournals which are available in the pharmaceutic industry. Thesereferences supply standard formulations which may be modified by theaddition or substitution of the reversible viscosifying composition ofthe present invention into the formulation. Suitable guidebooks includePharmaceutics and Toiletries Magazine, Vol. 111 (March, 1996);Formulary: Ideas for Personal Care; Croda, Inc, Parsippany, N.J. (1993);and Pharmaceuticon: Pharmaceutic Formulary, BASF, which are herebyincorporated in their entirety by reference.

The pharmaceutic composition may be in any form. Suitable forms will bedependant, in part, of the intended mode and location of application.Ophthalmic and otic formulations are preferably administered in dropletor liquid form; nasal formulations are preferable administered indroplet or spray form, or may be administered as a powder (as a snuff);vaginal and rectal formulations are preferably administered in the formof a cream, jelly or thick liquid; veterinary formulations may beadministered as a cream, lotion, spray or mousse (for application to furor exterior surface); esophageal and buccal/oral cavity applications arepreferably administered from solution or as a powder; film formingapplications or dermal applications may be administered as a lotions,creams, sticks, roll-ons formulations or pad-applied formulations.

Exemplary drugs or therapeutics delivery systems which may beadministered using the aqueous responsive composition compositions ofthe invention include, but are in no way limited to, mucosal therapies,such as esophageal, otic, rectal, buccal, oral, vaginal, and urologicalapplications; topical therapies, such as wound care, skin care and teatdips; and intravenous/subcutaneous therapies, such as intramuscular,intrabone (e.g., joints), spinal and subcutaneous therapies, tissuesupplementation, adhesion prevention and parenteral drug delivery. Inaddition, further applications include transdermal delivery and theformation of depots of drug following injection. It will be appreciatedthat the ionic nature of the biocompatible component of the responsivecomposition provides an adhesive interaction with mucosal tissue.

The responsive microgels of the present invention may be understood withreference to the following examples, which are provided for the purposesof illustration of example embodiments, and are not intended to limitthe scope of the claims appended hereto.

EXAMPLES Example I Drug Release from Responsive Microgels ofPolyether-Modified Poly(Acrylic Acid)

PLURONIC® F127 NF, L92 and L61 were obtained from BASF Corp. (MountOlive, N.J.) and used as received. The properties of these PLURONIC®surfactants are presented in Table 1: TABLE 1 Properties of thePLURONIC ® surfactants used in this study. M. Yu. Kozlov,Macromolecules, 2000, 33, 3305-3313; M. J. Kositza, et al., Langmuir,1999, 15, 322-325; BASF Catalog. Number Number Hydrophilic- Cloud pointCritical micelle Nominal of PO of EO lipophilic in water at 1concentration at Copolymer MW units units balance wt %, ° C. 25° C., ML61 2000 30 6 3 32 1.1 × 10⁻⁴ L92 3650 60 16 6 69 8.8 × 10⁻⁵ F127 1260065 200 22 >100 2.8 × 10⁻⁶

Materials

A fluorescent dye, 5-(4,6-dichlorotriazinyl)aminofluorescein (DCTAF,99%) was obtained from Molecular Probes, Inc. (Eugene, Oreg.). Acrylicacid (99%, vinyl monomer), ethylene glycol dimethacrylate (98%, divinylcross-linker, EGDMA), dodecane (99+%, solvent),4,4′-azobis(4-cyanovaleric acid) (75+%, azo initiator), and lauroylperoxide (97%, redox initiator) were purchased from Aldrich Chemical Co.and used as received. Poly(vinylpyrrolidinone-co-1-hexadecene) (GanexV-216) (dispersion stabilizer) was obtained from International SpecialtyProducts (Wayne, N.J.). Doxorubicin hydrochloride of 99% purity wasobtained from Hande Tech USA (Houston, Tex.), a subsidiary of YunnanHande Technological Development Co. (Kunming, P. R. China). All otherchemicals, gases and organic solvents of the highest purity availablewere obtained from commercial sources.

Microgel Synthesis

Synthesis was carried out on a laboratory scale in an adiabatic mode.Acrylic acid (vinyl monomer) (40 mL) was partially neutralized byaddition of 5 M NaOH aqueous solution (0.5 mL). PLURONIC® F127 or L92(24 g) was dissolved in the resulting solution under nitrogen and adesired amount of ethylene glycol dimethacrylate (EGDMA) (divinylcross-linker) was added. Amounts of EGDMA were set such that resulted in1 mol % relative degree of cross-linking of the microgels [cross-linkingmol %=100×(number of mols of EGDMA/the number of mols of acrylic acid)].Lauroyl peroxide (redox initiator) (100 mg) and4,4′-azobis(4-cyanovaleric acid) (azo initiator) (100 mg) were dissolvedin 2 mL of acrylic acid (vinyl monomer) and added to the solution ofPLURONIC® in acrylic acid. The resulting solution was deaerated bynitrogen bubbling for 0.5 h and added to a 3-necked 0.5-mL flaskcontaining 1 wt % solution of Ganex V-216 (dispersion stabilizer) indodecane (solvent) (200 mL). The flask was vigorously stirred by amechanical stirrer and deaerated by constant nitrogen purge from thebottom. Then the flask was heated to 70° C. using an oil bath and keptat that temperature under stirring and nitrogen purge. After about 1 h,formation of white particles was observed on the flask walls. Thereaction was continued at 70° C. for another 3 h. Then the reactor wasdisassembled, and the contents of the reactor were filtered usingWhatman filter paper (retention size 10 μm). The microgel particles wereextensively washed by hexane and dried under vacuum. Spherical particleswere observed under microscope. Particle sizing was performed in hexaneusing a ZetaPlus Zeta Potential Analyzer (Brookhaven Instruments Co.).Typical batches of particles made with PLURONIC® F127 and L92 weremeasured to have effective median diameter of 13 μm (polydispersity 1.4)and 6 μm (polydispersity 1.3), respectively.

In order to ascertain the grafting of PLURONIC® segments ontocross-linked poly(acrylic acid) [poly(vinyl monomer)] networks, aparticulate sample was suspended in 1 M NaOH for 3 days and lyophilized.The sample was then placed into a Soxhlet extractor charged withdichloromethane and kept under reflux for 2 days. The wash-outs werecollected, evaporated under vacuum, and weighed. Preliminary experimentsdemonstrated negligible solubility of poly(sodium acrylate), and totalsolubility of the PLURONIC®, respectively, in dichloromethane. Thefraction of the PLURONIC® washed from the particles was negligible,within experimental error (±5% of the initial PLURONIC® content). Hence,for all practical purposes, the overall composition of the microgels inthe present study corresponded to that set in the reactor. That is, theweight ratio of PLURONIC® to poly(acrylic acid) [poly(vinyl monomer)] inthe particles was 43:57.

Synthesis of Labeled PLURONIC® L61

The DCTAF-labeled PLURONIC® L61 was synthesized and purified essentiallyas described by Ahmed, F., et al., S. Langmuir, 2001, 17, 537-546. Stocksolutions of 6 w/v % PLURONIC® L61 were prepared by dissolving thepolymer in 0.1 M sodium bicarbonate solution at pH=9.30. A stocksolution of 20 g/L 5-DTAF was prepared by dissolving the fluoresceinprobe in dimethyl sulfoxide (DMSO). The 5-DTAF solution was diluted in0.1 M sodium bicarbonate solution and added to the PLURONIC® blockcopolymer solution such that the molar ratio of 5-DTAF to PLURONIC® was1:1. The reaction was allowed to proceed in the dark at room temperatureovernight. To separate the labeled PLURONIC® from the excess unreacted5-DTAF, the size exclusion chromatography was applied. Sephadex G-25beads (Aldrich Chemical Co.) swollen in boiling 0.05 M NaCl solutionwere first packed into a Chromaflex column (ID, 2.5 cm, length, 60 cm)(Kimble/Kontes, Vineland, N.J.). The column was then primed by washingwith 0.05 M NaCl solution, followed by passage of 1 bed volume of 6 w/v% unlabeled PLURONIC® in sodium bicarbonate solution and 2-3 bed volumesof 0.05 M NaCl solution. A 250 μL sample of the reaction mixture wasthen added to the column, and the labeled product was eluted with NaClsolution. The yellow bands moving down the length of the column wereseparated and the PLURONIC®-containing fraction was concentrated using aCentricon centrifugal filter device (Millipore Corp., Bedford, Mass.)with a molecular weight cutoff of 1500. The samples were centrifuged atca. 5000×g for 1 h. The retentate containing labeled PLURONIC® waslyophilized and kept at −20° C. in the dark. Given the dye molarextinction coefficient of 83000 M⁻¹ cm⁻¹ and molecular weight of 495.3,the efficiency of the DCTAF dye conjugation with PLURONIC® molecule wasestimated to be 2.4-3.0%.

Solute Loading onto Microgels

The loading level of doxorubicin or PLURONIC® L61 labeled with DCTAFinto microgels was measured using a Millipore Ultrafree-MC CentrifugalFilter Device (Millipore Corp.). A microgel was suspended in Tris buffer(10 mM, pH 7.0) and 1 mL of the suspension (40 mg gel/mL buffer) wereequilibrated with 6.0 mM stock solution of a drug (9 mL) for 72 h whileshaking. Shaking was performed using a KS10 orbital shaker(BEA-Enprotech Corp., Hyde Park, Mass.) in an environmental chamber at37° C. After equilibration, the microgel particles were filtered off bycentrifugation (10000×g, 0.5 h) and supernatant was assayed for drugconcentration. A Shimadzu Model 1600 PC spectrophotometer with atemperature-controlled quartz cuvette (path length 1 cm) was used forelectronic absorption measurements. The extinction coefficients ofdoxorubicin (λ=482 nm) was determined to be 12300 M⁻¹ cm⁻¹. Assuming theaverage molecular weight of 2000, the extinction coefficients of theDCTAF-labeled PLURONIC® (λ=492 nm) was measured at pH 7.0 to be 2600 M⁻¹cm⁻¹. The drug uptake was calculated from the absorbance readings in theappropriately diluted stock solution and in the system equilibrated withmicrogel. The U values were measured in triplicate for each drug andgel, respectively. In a control series of experiments, equilibration of24 μmol/mL doxorubicin with microgels for 1 week yielded U values close(within experimental error) to the ones obtained with 6 μmol/mLsolutions under otherwise identical conditions (see above). This ensuredequilibrium U values. The gels equilibrated with corresponding drugswere snap-frozen in liquid nitrogen, lyophilized, and stored at −70° C.in the dark. In the subsequent release studies, the dry gel powders ofknown U were reconstituted with Tris buffer (10 mM, pH 7.0) to result inthe known concentration of the gel and drug.

Release Studies

Drug release from microgels loaded with either DCTAF-labeled PLURONIC®or doxorubicin was studied using the dynamic dialysis technique ofGupta, P. K., et al., J. Pharm. Sci., 1987, 76, 141-145. A Teflon-made,thermostatted dynamic dialyzer consisted of two chambers, separated by adialysis membrane (cellulose ester, working area A=0.35 cm², molecularweight cut-off 100 kDa, Spectrum Laboratories). A cylindrical feedchamber (volume V_(f)=5.0 mL) containing drugs or drug-loaded gels wasvigorously stirred by a magnetic bar, while a receiver chamber had aninlet and outlet for a constant flow of the receiver solution. Thereceiver solution (Tris buffer, 10 mM, pH 7.0) was circulated along thedialysis membrane using a P625/275 peristaltic pump (InstechLaboratories). Concentration of the drug in the receiver solution wasmonitored online by passing through a thermostatted quartz cuvette (pathlength 1.0 cm). Concentration of solutes was measured periodically usinga Shimadzu Model RF-5301 PC spectrofluorophotometer (slit widths 3.0nm). The dialyzer was maintained at 37° C. by submersion in a waterbath. The flow rate through the receiver chamber was maintained at 1.5mL/min. In a series of preliminary experiments, it was established thatat this flow rate the doxorubicin transport becomes flowrate-independent, and yet the flow of the PLURONIC® solution affordsavoiding any foaming. The total volume of the receiver solution (V_(r)),including the chamber, cuvette, and 0.093″ tubing, was 98 mL in allexperiments.

Permeability of the dialysis membrane was determined by loading acertain amount (q_(f) ⁰) of either doxorubicin or PLURONIC® solutioninto the feed chamber and measuring the kinetics of release. Thedialysis membrane had been soaked in the corresponding solute solutionfor 48 h prior to the kinetic measurement. To remove excess solute, themembrane was gently wiped up by a paper tissue on both sides immediatelyprior to the loading into the dialyzer. The membrane thickness (δ) wasmeasured microscopically in the receiver solution upon completion of thedialysis experiment and was typically in the order of 100 μm. Solvingthe 1^(st) Fick's law expression for the diffusion across the dialysismembrane $\begin{matrix}{{Flux} = {\frac{\mathbb{d}q_{r}}{\mathbb{d}t} = {D\frac{A\left( {C_{f} - C_{r}} \right)}{\delta}}}} & (1)\end{matrix}$yields an expression that allows for an estimation of the apparentdiffusion coefficient (D):ln └q_(f) ⁰ −C _(r)(V _(f) +V _(r))=lnq _(f) ⁰ −Kt  (2)where K=DA(V _(f) +V _(r))/V _(f) V _(r)δ  (3)Herein, q(t) and C(t) are the drug quantity and concentration,respectively, and subscripts f and r designate the feed and receiversolution, respectively.

Having defined the ranges of concentrations where the diffusioncoefficient of either doxorubicin or PLURONIC® was independent of thedrug initial concentration C_(f) ⁰, we used the microgel loading thatdid not exceed these ranges. A known mass of microgel particles with aknown loading (U, see above) suspended in 10 mM Tris buffer (pH 7.0) wasplaced in the feed chamber resulting in an initial drug quantity in thesystem, Q_(o). Assuming that the drug decay in the microgel particlescan be approximated by a single-exponential (i.e., first-order) kineticsQ(t)=Q_(o)e^(−K) ¹ ^(t), the Fick's law expression (1) can be rewrittenas $\begin{matrix}{{Flux} = {\frac{\mathbb{d}C_{r}}{\mathbb{d}t} = {\frac{DA}{\delta\quad V_{r}}\left\lbrack {{\frac{Q_{0}}{V_{f}}\left( {1 - {\mathbb{e}}^{{- K_{1}}t}} \right)} - {C_{r}\left( {\frac{V_{r}}{V_{f}} + 1} \right)}} \right\rbrack}}} & (4)\end{matrix}$and solved with respect to C_(r) as follows: $\begin{matrix}\begin{matrix}{C_{r} = {\frac{Q_{0}}{V_{f} + V_{r}}\left\lbrack {1 - {\mathbb{e}}^{- {Kt}} - {\frac{{DA}\left( {V_{f} + V_{r}} \right)}{\delta\quad V_{f}{V_{r}\left( {K - K_{1}} \right)}}{\mathbb{e}}^{- {Kt}}} -} \right.}} \\\left. {\frac{{DA}\left( {V_{f} + V_{r}} \right)}{\delta\quad V_{f}{V_{r}\left( {K - K_{1}} \right)}}{\mathbb{e}}^{{- K_{1}}t}} \right\rbrack\end{matrix} & (5)\end{matrix}$

At long dialysis times and Kt>>K₁t, eq(5) can be simplified:$\begin{matrix}{{\ln\left\lbrack {\frac{Q_{0}}{V_{f} + V_{r}} - C_{r}} \right\rbrack} = {{\ln\left\lbrack \frac{{DAQ}_{0}}{\delta\quad V_{f}{V_{r}\left( {K - K_{1}} \right)}} \right\rbrack} - {K_{1}t}}} & (6)\end{matrix}$Equation (6) indicates that a plot of ln [Q_(o)/(V_(f)+V_(r))−C_(r)] vstime should be a straight line that yields K₁. We used equations (2) and(3) to calculate the permeability of the dialysis membrane inexperiments without microgels, and equation (6) to estimate K₁ inexperiments with drug-loaded microgels.

Results

Kinetics of the release of doxorubicin and PLURONIC® L61 throughdialysis membrane with and without microgels are compared in FIG. 5(Kinetics of cumulative release of doxorubicin (circles) and PLURONIC®L61 (triangles) through the dialysis membrane with (filled points) andwithout (open points) microgels in the feed chamber at 37° C. Receiverchamber comprised 10 mM Tris buffer (pH 7.0) in all cases. Initialdoxorubicin concentration in the feed 110 μg/mL (open circles) and 20mg/mL (filled circles), initial PLURONIC® L61 concentration in the feed20 mg/mL (open triangles) and 22 mg/mL (filled triangles). Microgelsused in the feed were composed of PLURONIC® F127 and poly(acrylicacid)).

As shown in FIG. 5, loading of the corresponding drugs into microgelsaffected the kinetics of release greatly. Without microgels, almost 100%of the drug was released within less than a day, while the drugs loadedinto microgels exhibited slow, sustained release kinetics.

In the control experiments, we measured permeability of the dialysismembrane for doxorubicin and PLURONIC® L61 without loading intomicrogels. A range of initial drug concentrations in the feed solutionswas explored, in order to verify the independence of the effectivemembrane permeability of the q_(f) ⁰, as is required if the change ofthe solute size (i.e. aggregation and/or micellization) is absent. FIG.6 illustrates kinetics of the drug diffusion through the dialysismembrane as functions of the initial drug concentrations in the feed.The effective membrane permeability constant, K, was found from theslopes of the corresponding linear fits using equation (2). Thusobtained K values were equal to 0.70±0.048(4) and 0.41±0.019(3) h⁻¹ μg⁻¹for doxorubicin and PLURONIC® L61, respectively. Given very close Kvalues (standard error below 7% and 5% for doxorubicin and PLURONIC®L61, respectively) obtained within the studied ranges of C_(f) ⁰ weconcluded that aggregation was absent within those ranges. Using mean Kand measured membrane thickness (δ) values, we obtained the effectivediffusion coefficients (D) of 2.4×10⁻⁵ and 1.4×10⁻⁵ cm²/s fordoxorubicin and PLURONIC® L61, respectively (eqn (3)). These values arereasonably close to D of the corresponding solutes in water, indicatingthat the chosen dialysis membrane does not constitute any significantdiffusional barrier to these solutes in their non-aggregated state. FIG.6 shows the kinetics of doxorubicin and PLURONIC® L61 release throughthe dialysis membrane from an aqueous feed solution at 37° C., expressedin terms of equation (2). Numbers stand for C_(f) ⁰, μg/mL.Corresponding linear fits (R²>0.99 in all cases) were used to calculate(eqn(2)) the effective membrane permeability constant, K.

Having defined the membrane permeability, we proceeded to the drugrelease study from the microgels. Kinetics of doxorubicin release fromthe microgels are shown in FIG. 7. The effective release constants (K₁)for doxorubicin were measured to be (14.2±0.36)×10⁻³ and(22.8±0.49)×10⁻³ h⁻¹ μg⁻¹ for microgels based on PLURONIC®F127-PAA-EGDMA and PLURONIC® L92-PAA-EGDMA, respectively. FIG. 7 showsthe kinetics of doxorubicin release through the dialysis membrane frommicrogels at 37° C., expressed in terms of equation (6). Numbers standfor C_(f) ⁰ in μg/mL. The datapoints were fitted to linear fits (R²>0.98in all cases) with the slopes used to calculate (eqn(6)) the effectiverelease constant, K₁. In A, the gels used in feed consisted of PLURONIC®F127 and PAA, whereas in B, the gels consisted of PLURONIC® L92 and PAA.The effective degree of the gel cross-linking was 1 mol % throughout.

Kinetics of PLURONIC® L61 release are presented in FIG. 8 which showsthe release through the dialysis membrane from microgels at 37° C.,expressed in terms of equation (6). Numbers stand for C_(f) ⁰ in μg/mL.The datapoints were fitted to linear fits (R²>0.97 in all cases) withthe slopes used to calculate (eqn(6)) the effective release constant,K₁. In A, the gels used in feed consisted of PLURONIC® F127 and PAA,whereas in B, the gels consisted of PLURONIC® L92 and PAA. The effectivedegree of the gel cross-linking was 1 mol % throughout.

The effective release constants (K₁) for the PLURONIC® were measured tobe (2.1±0.18)×10⁻³ and (0.44±0.046)×10⁻³ h⁻¹ μg⁻¹ for microgels based onPLURONIC® F127-PAA-EGDMA and PLURONIC® L92-PAA-EGDMA, respectively. FIG.8 shows that a very slow, sustained release of the PLURONIC L61® wasachieved within at least 10 days, with cumulative concentrations reachedin the receiver solution (C_(r)) that did not exceed 10-14% of theinitial loading, C_(f) ⁰. The exceptionally low release rate of thePLURONIC® L61 can be explained by the formation of mixed micellesbetween added PLURONIC® L61 and PLURONIC® covalently grafted to the PAAnetwork in the process of synthesis. Such mixed, immobile micelles canprovide thermodynamically stable environment for the PLURONIC® solute,making its effective partition coefficient between micelles and water tobe very low. This notion is supported by the observation that therelease rate from the gels from L92-PAA-EGDMA was about 5-fold higherthan from the gels containing PLURONIC® F127 bonded to PAA. Formation ofstable mixed micelles between relatively hydrophobic PLURONIC® L61 andL92 can be favored than between PLURONIC® L61 and relatively hydrophilicF127 (for PLURONIC® properties, see Table 1, supra).

Example II Microgel Synthesis

Nonionic copolymer PLURONIC® F127 NF was obtained from BASF Corp. andused without further treatment. It has a formula EO₁₀₀PO₆₅EO₁₀₀, nominalmolecular weight 12600, molecular weight of PPO segment 3780, 70 wt % ofEO, and cloud point above 100oC. Acrylic acid (99%) (vinyl monomer),ethylene glycol dimethacrylate (EGDMA) (98%) (divinyl cross-linker),dodecane (99+%) (solvent), and 4,4′-azobis(4-cyanovaleric acid) (75+%)(azo initiator) were purchased from Aldrich Chemical Co. and used asreceived. Lauroyl peroxide (97%) (redox initiator) was obtained fromFluka Chemie AG, Switzerland. Poly(vinylpyrrolidinone-co-1-hexadecene)(Ganex V-216) (dispersion stabilizer) was obtained from InternationalSpecialty Products and used as received. All other chemicals, gases andorganic solvents of the highest purity available were obtained fromcommercial sources.

Synthesis was carried out on a laboratory scale in an adiabatic mode.Acrylic acid (vinyl monomer) (40 mL) was partially neutralized byaddition of 5M NaOH aqueous solution (0.5 mL). PLURONIC® F127 NF (23.4g) was dissolved in the resulting solution under nitrogen and a desiredamount of ethylene glycol dimethacrylate (EGDMA) (divinyl cross-linker)was added. Amounts of EGDMA ranged from 1.1 μL to 1.1 mL and the molarratio of the EGDMA to acrylic acid set in the reaction mixturedesignates the degree of cross-linking of the microgels in what follows.Lauroyl peroxide (100 mg) and 4,4′-azobis(4-cyanovaleric acid) (100 mg)were dissolved in 2 mL of acrylic acid and added to the solution ofPLURONIC® F127 NF in acrylic acid. The resulting solution was deaeratedby nitrogen bubbling for 0.5 h and added to a 3-necked 0.5-mL flaskcontaining 1 wt % solution of Ganex V-216 in dodecane (200 mL). Theflask was vigorously stirred by a mechanical stirrer and deaerated byconstant nitrogen purge from the bottom. Then the flask was heated to70° C. using an oil bath and kept at that temperature under stirring andnitrogen purge. After about 1 h, formation of white particles wasobserved on the flask walls. The reaction was continued at 70° C. foranother 3 h. Then the reactor was disassembled, and the contents of thereactor were filtered using Whatman filter paper (retention size 10micron). The microgel particles were extensively washed by hexane anddried under vacuum.

Spherical particles were observed under microscope. Particle sizing wasperformed in hexane using a ZetaPlus Zeta Potential Analyzer (BrookhavenInstruments Co.). A typical batch containing 1 mol % cross-linking wasmeasured to have effective

Example III Microgel Superabsorbent Properties

The ability of microgels to absorb water was estimated using avolumetric method. Single microgel particles were placed into glasscapillary tubes (internal diameter 1-1.2 mm) using suction pressuresapplied by an Ultramicro Accropet filler/dispenser via rubber connector.The tubes were placed into a homemade glass thermostatted cuvet andobserved under an inverted microscope equipped with a microscaler and avideo monitor. Similar experimental setup was described in Eichenbaum,G. M., et al., Macromolecules, 1998, 31, 5084-5093. The boundaries ofthe spherical particles were fitted with the microscaler and a particlediameter was measured with an accuracy of ±0.5 μm or better. Initially,a diameter of a dry particle (d_(o)) was measured, then the capillarytube was gently filled with deionized water (pH adjusted by 5 M NaOH)immersed into a reservoir of the same solution. The diameter of theswollen particle (d_(s)) was measured at a given temperature. Theparticles were allowed to swell for 24 h, after which no changes in theparticle size were observed at any temperature. The equilibrium volumeratio S=V/V_(o) was defined as S=(d_(s)/d_(o))³. Measurements at givenpH and temperature were conducted with 5 different particles indifferent capillary tubes. Average S values are reported throughout.

The results of equilibrium swelling experiments with microgelscross-linked by EGDMA are shown in FIG. 9. The length of subchain (i.e.length of the chain between cross-links) N defined as in Bromberg, L.,et al., J. Chem. Phys., 1997, 106, 2906-2910.N=[a ⁶ c _(xl)(c _(xl) +c _(m))]⁻¹where a=10v_(xl)v_(m), v_(xl)=0.2 M⁻¹ is the molar volume of thecross-linker (EGDMA), and v_(m)=0.063 M⁻¹ is the molar volume of themonomer (acrylic acid).

FIG. 9. Equilibrium swelling of microgel particles in deionized water asa function of the length of subchain. pH 7.0. The results shown in FIG.9 indicate that at 15° C., swelling of the microgels corresponds to theswelling of other superabsorbents and is governed by elasticity of thepermanent cross-links and osmotic term corresponding to theelectrostatic repulsion of the chains. At 15° C., the swelling ratio Sscales as S∝N^(0.6), which according to the Flory-Huggins theory isindicative of the Gaussian chain statistics, typical for covalentlycross-linked gels. However, at 37° C., elasticity of the microgelbecomes higher due to the appearance of additional cross-links(PLURONIC® chains with hydrophobic poly(ethylene oxide) segments). Theswelling ratio scales as S∝N^(x), with x<0.6 meaning non-Gaussian chainstatistics. Overall, the results in FIG. 9 show i) high absorbency(swelling ratio in DI water up to 300 and higher), and ii) usefultemperature sensitivity of water uptake.

Example IV pH-Sensitivity of Microgel Swelling

Experiments were conducted as described in Example III, except themicrogel particles were allowed to equilibrium swell at a certain pH_(o)and temperature to yield a d_(o). Then the aqueous solution was gentlyremoved from the glass tube by filter paper and the tube with themicrogel particle was immersed into a solution of different pH_(x) andequilibrated there overnight. Finally, the tube with the microgelparticle filled with the solution of pH_(x) was inserted into the cuvetand thermostatted at desired temperature to yield an equilibriummicrogel diameter d_(x). The equilibrium volume ratio S=V_(x)/V_(o) wasdefined as S=(d_(x)/d_(o))³. FIG. 10 shows quilibrium swelling ofmicrogel particles in deionized water at 15° and 37° C. as a function ofpH. Degree of cross-linking in molar percent is indicated. Results inFIG. 10 show dramatic increase in swelling above pH 3.8-4.1,corresponding to pKa of poly(acrylic acid). Hence, our microgels wouldbe collapsed at pH 1-2 and fully swollen at pH 7.4. This is a usefulproperty applicable in oral or colonic drug delivery.

Example V Temperature-Sensitivity of Microgel Swelling

Experiments were conducted as described in Example IV and the resultsare shown in FIG. 11 illustrating equilibrium swelling of microgelparticles in deionized water at pH 7.0 as a function of temperature.Degree of cross-linking in molar percent is indicated. These resultsdemonstrate useful temperature-sensitivity of the microgel swelling.

Example VI Temperature-Sensitivity of Solubilization of HydrophobicCompounds

Solubilization of pyrene, a well-known hydrophobic fluorescent probe,was used to reveal formation of aggregates within microgel particlescapable of solubilizing hydrophobic compounds. The microgel particleswere suspended in DI water and pH of the suspension was adjusted to 7.0using 10 M NaOH. A stock solution of 1 mM pyrene in absolute methanolwas prepared, from which 1-3 μL were added to an aerated 1 wt %suspension resulting in 0.6 μM pyrene concentration. The sample was thenallowed to equilibrate for 20 min at a given temperature and emission(μ_(ex)=335 nm) spectra were recorded using a stirred, thermostattedquartz cell with a 1-cm path length. The spectra were measured undercontrolled temperature conditions using a Shimadzu Model RF-5301 PCspectrofluorophotometer (slit widths of 1.5 nm). The ratio of theintensities of the first (373 nm) to the third (384 nm) vibronic peak(I₁/I₃) in the emission spectra of the monomer pyrene were used toestimate the polarity of the pyrene microenvironment. For comparison, 1wt % solutions of PLURONIC® F 127 NF were prepared and studied in thesame fashion. The effect of temperature on I₁/I₃ of pyrene in microgelsuspension (1 mol % cross-linking) or in polymer solutions is presentedin FIG. 12. FIG. 12 shows the effect of temperature on the ratio of thefirst-to-the-third vibronic band intensities (I₁/I₃) of pyrene in 1 wt %microgel suspension or in 1 wt % PLURONIC® F127 solution. Microgel with1 mol % cross-linking is designated F127-PAA-EGDMA. pH 7.0 throughout.As is seen, in 1.0% PLURONIC® F127 solutions, for example, the I₁/I₃sharply declines above 20° C., which is the critical micellizationtemperature (CMT). Alexandridis, P., et al., J. Am. Oil Chem. Soc. 1995,72, 823. At temperatures below CMT, the I₁/I₃ is only slightly below theI1/I3 in water, indicating high polarity of the pyrene environment andlack of solubilization. The microgel suspension has I1/I3 values thatare significantly less than in the corresponding Pluronic solution,indicating lesser polarity and higher capability of solubilizationthroughout the temperature range. The I1/I3 in microgel suspension islow at T<20° C. (which corresponds to the critical micellizationtemperature of PLURONIC®) and increases in the temperature range 20-26°C. Above 26° C., the I₁/I₃ decreases. At low temperatures, hydrophobicdomains exist in the microgels that are getting solubilized intohydrophobic micelle-like aggregates of Pluronic within microgels. Oncemicelles are formed above 26° C., they provide a hydrophobic environmentfor the pyrene. The temperature-responsive mode of solute solubilizationby microgels is useful for medicinal and cosmetic formulations.

Example VII Loading of Ionic and Hydrophobic Drugs

Water-Soluble Solutes

The maximum loading level of doxorubicin, mitoxantrone, and mitomycin C,for example, into microgels was measured using a Millipore Ultrafree-MCCentrifugal Filter Device (Millipore, Co.). A microgel was suspended inTris buffer (5 mM, pH 7.0) and 50 μL of the suspension (2 mg gel/mLbuffer) was equilibrated with 3.0 mM stock solution of a drug (450 μL)for 16 h while shaking 44, 45. Shaking was performed using a KS 10orbital shaker (BEA-Enprotech Corp., Hyde Park, Mass.) in anenvironmental chamber at 37° C. In the case of doxorubicin, pH of themicrogel suspensions equilibrated with the stock drug solution wasvaried by addition of small amounts of 5 M NaOH or HCl solutions, andtemperature was varied from 15 to 45° C. After equilibration, themicrogel particles were filtered off by centrifugation (10000×g, 0.5 h)and supernatant was assayed for drug concentration. A Shimadzu Model1600 PC spectrophotometer with a temperature-controlled quartz quvette(path length 1 cm) was used for electronic absorption measurements. Theextinction coefficients of doxorubicin (λ=482 nm) and mitoxantrone(λ=614 nm) were determined at pH 7.0 to be 12200 and 22100 M⁻¹ cm⁻¹,respectively. The concentration of mitomycin C was assayed by HPLC usinga Capcell Pak MF Ph-1 (100×4.6 mm I.D., particle size 5 μm) column(Phenomenex, Torrance, Calif.). The HPLC was a Hewlett-Packard 1090system with an autosampler and a variable wavelength UV detectorcontrolled by the HPLC Chemstation software (Hewlett-Packard). Deionizedwater was used as a mobile phase (flow rate, 1 mL/min, injection volume,25 μL), and detection was carried out at 365 nm 47. Typical retentiontime of the mitomycin C was 4.88 min.

The drug uptake was expressed as:

U (mmol drug/g gel)=[(Ac−Ar)/Ac]VCs/Mgel, where Ac and Ar are theabsorbance or HPLC readings in the appropriately diluted stock solutionand in the system equilibrated with microgel, respectively, V=0.5 ml isthe total volume of the system, Cs=3 μmol/mL is the concentration of thestock solution, and Mgel=0.1 mg is the microgel mass. The U values weremeasured in triplicate for each drug and for each temperature, pH, andgel, respectively. In a control series of experiments, equilibration of6 μmol/mL doxorubicin with microgels for 1 week yielded U values close(within experimental error) to the ones obtained with 3 μmol/mLsolutions under otherwise identical conditions. This ensured equilibriumU values.

Hydrophobic Solutes

The loading of taxol into microgels was measured by equilibrating taxoladsorbed onto steel beads with the 1 wt % suspension of microgels (pH7.0). Stainless steel beads (1-3 mm diameter) were soaked in 10 mMsolution of taxol in acetonitrile, following by stripping off thesolvent in a rotary evaporator. The beads were used in order to enhancethe area of contact between microgel suspension and taxol. The beadswere separated into several fractions. One fraction was added to apolypropylene vial containing the microgel suspension (0.5 mL) and thevial was gently shaken in a horizontal position in an environmentalchamber at 20 or 37° C. Then the beads were recovered from thesuspension by using a magnet. The beads were then dried under vacuum andplaced into acetonitrile (0.5 mL), where taxol was extracted aftershaking overnight. The solvent fraction was assayed for taxolconcentration using HPLC. The control fraction of loaded beads wassubjected to the extraction without equilibration with the microgelsuspension. The solubility of taxol in water was measured at 37° C. bysonicating 5 mg of the drug suspension in 0.5 ml water placed in apolypropylene vial for 15 s followed by centrifugation at 10000 g for 3min. The supernatant was then removed, evaporated under vacuum, thetaxol traces were dissolved in acetonitrile and assayed by HPLC. Taxolconcentrations were measured in triplicate using HPLC system describedabove. The chromatography assay comprised the use of a Capcell Pak C18UG 120 (150×4.6 mm I.D., particle size 3 μm) column (Phenomenex),acetonitrile-0.1% phosphoric acid in DI water (55:45 v/v, 1.3 mL/min) asa mobile phase, and UV detection at 227 nm.

Typical retention time of the taxol peak was 3.46 min.

Results

Three cationic and one uncharged drugs were loaded onto the microgels.All of these compounds are currently in clinical use as anticancerdrugs. Doxorubicin, mitoxantrone, and mitomycin C are mono-, di-, andtrivalent cationic weak bases, respectively. FIG. 13 shows theequilibrium uptake of doxorubicin by microgels (crosslinking (XL)=1 mol%) as a function of pH at 37° C.

Table 2 lists molecular weights, n-octanol-to-water partitioncoefficients (P), and equilibrium uptake of the drugs into the microgelscharacterized by XL=1 mol % and maximum ion-exchange capacity of 6.12mmol/g (measured by bulk titration as described herein). All of theuptake values in Table 2 were less than or equal to maximum microgelcapacity for protons. A pronounced dependence was observed with the weakbases: the smaller, more hydrophilic, and more charged solutes had thehigher loading into the microgels. TABLE 2 Properties of anticancerdrugs and their equilibrium uptake by the PLURONIC ®-PAA microgels(cross-linking ratio, XL = 1 mol %, ion-exchange capacity, 6.12 mmol/g)at pH 7.0. Drug MW ^(a) Log P Charge Uptake ± SD, mmol/g Mitomycin C334.1 −0.4  3 5.31 ± 1.86 (37° C.) Mitoxantrone 444.2   0.24 ^(b) 2 3.70± 0.56 (37° C.) Doxorubicin 543.5 1.85 1 2.97 ± 0.33 (20° C.) 2.26 ±0.37 (37° C.) Taxol 853.3 4   0 (2.27 ± 0.90) × 10′⁻³ (20° C.) (6.97 ±0.87) × 10′⁻³ (37° C.)^(a) Molecular weights are given for free bases, and not hydrochloridesalts.^(b) Calculated using ClogP Program.

The characteristic increase in taxol loading capacity at temperaturesabove CMT provides additional evidence to the mechanism of taxolsolubilization into micelle-like aggregates within microgels. Themicelles in PLURONIC®-PAA solutions typically have solubilizing capacityhigher than the small hydrophobic domains existing below CMT. Thesolubilizing capacity of the microgels for taxol is at least equal tothat of PLURONIC®-PAA micelles for other hydrophobic solutes such assteroid hormones. The ability of microgels to effectively load and holdtaxol, combined with mucoadhesive properties is a feature important forlocalized delivery.

General trends important for drug loading via ion-exchange mechanismwere studied using the potent chemotherapeutic drug doxorubicin. As thedegree of carboxyl group ionization increases with pH, the ion-exchangecapacity of the microgels increase, reaching about half of the maximumcapacity found by titration, indicating that the loading of doxorubicincan be limited by the available free volume of the network. Notably, thepH-dependencies of the equilibrium swelling and doxorubicin loadingcoincide. Similar result was observed with poly(methacrylic acid)microgels. The effects of steric “crowding” of the drug within themicrogel network and the availability of the carboxyls for theion-exchange are highlighted by the effects of temperature andcross-linking density. The collapse of the microgels at elevatedtemperature due to the appearance of physical cross-links leads tolesser volume of the microgel network available for hosting the drug,and thus lower equilibrium loading at higher temperatures. Analogously,the longer subchain allowing for the looser network and higher swellingleads to the higher equilibrium loading of doxorubicin. The very highoverall capacity of the microgels for doxorubicin (2 M and higher), willallow proper chemotherapeutic drug delivery. Microgels loaded with bothtaxol and doxorubicin, for example, is a feature embodiment of themicrogels described herein.

Example VIII Doxorubicin Transport Study Across Gastrointestinal Caco-2Layers Materials and Cell Culture

Caco-2 cells (American Type Culture Collection, Rockville, Md.) weremaintained at 37° in Dulbecco's Modified Eagle Medium (DMEM) containingN-(2-hydroxyethyl)piperazine-N′-(2-ethanesulfonic acid) (HEPES, 25 mM),glucose (4.5 g/L), and supplemented with 10% (v/v) fetal bovine serum,1% nonessential amino acids, L-glutamine (2 mM), penicillin (100000U/L), and streptomycin (100 mg/L), in an atmosphere of 10% CO₂ and 90%relative humidity. The cell line (passage numbers from 70 to 85) wassubcultured by trypsinization every week and the medium was replacedevery other week. Cells were passaged at 90% confluency using a 0.25%trypsin/0.20% ethylene diamine tetraacetic acid (EDTA) solution. Allcell culture products were received from GIBCO™ (Invitrogen Corporation,Carlsbad, Calif.). Hank's balanced salt solution (HBSS, composition:KH₂PO₄, 0.44 mM; KCl, 5.37 mM; Na₂HPO₄, 0.34 mM; NaCl, 136.9 mM;D-glucose 5.55 mM) buffered with 30 mM HEPES at pH 7.2,3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, andVerapamil were obtained from Sigma Chemical Co. (St. Louis, Mo.).D-[1-¹⁴C]-mannitol (250 mCi/mmol, 99+% radiochemical purity, MW 182.2)was obtained from PerkinElmer Life Sciences (Boston, Mass.). Carbopol®934 NF polymer was received from Noveon, Inc. (Cleveland, Ohio).Microgels comprising copolymers of poly(acrylic acid) and Pluronic F127NF or Pluronic L92 cross-linked by ethylene glycol dimethacrylate ateffective cross-linking density of 0.1 mol % were synthesized by afree-radical suspension polymerization as described previously [9]. Themicrogels were purified from admixtures of acrylic acid and unattachedPluronic by Soxhlet extraction and dialysis [9]. The purified microgelswere analyzed for residual unattached Pluronic as follows. A microgelsuspension (10 wt %, 50 g) in deionized water was dialyzed for a week at4° C. against deionized water (0.5 L) using a Spectra/Por® celluloseester membrane (molecular weight cut-off 100,000, Spectrum®, LagunaHills, Calif.). Then the dialysis bag was withdrawn and the dialyzatewas lyophilized to dryness in 40-mL polypropylene tubes. Then the tubeswere rinsed, with brief sonication, with total of 5 mLdimethylformamide, thereby concentrating the wash-outs, if any,100-fold. The wash-outs were analyzed for the presence of Pluronic usingHPLC. Liquid chromatography was performed using a Hewlett-Packard model1050 chromatograph, a Dynamax model RI-1 analytical refractive indexdetector, and two PLgel 5 μm mixed ‘D’ columns in series. The analysesof the residual solids were run in DMF at 1.0 mL/min flow rate usingpoly(ethylene glycol) standards kit (Polysciences, Inc., Warrington,Pa.) for molecular weight calibration. The eluent was DMF (HPLC grade)at a flow rate of 1 mL min⁻¹. In the control assays, a 20 μg/mLconcentration or lower of either Pluronic F127 or L92 could be detected.However, no Pluronic could be detected in the wash-outs, which meansthat the concentration of the unattached Pluronic was less than 0.01 wt% of the total Pluronic bonded onto microgels. The weight ratio ofPluronic to poly(acrylic acid) in purified microgels used in this studywas determined as described elsewhere [7] to be 45:55. The microgelsequilibrium swollen in deionized water (pH adjusted to 7.0) weresubjected to particle sizing using an AccuSizer 780/SPOS (ChristisonScientific Equipment Ltd., Gateshead, UK). The mean populations of theF127-PAA-EGDMA and L92-PAA-EGDMA microgels were of 54±22 and 23±9 μmsize, respectively.

Transepithelial Transport of Doxorubicin

The drug-containing samples for the transport experiments were preparedas follows. A freeze-dried microgel was autoclaved at 121° C. for 15 minand suspended in serum-free DMEM, where it was allowed to equilibrate at37° C. under gentle stirring overnight in sterile conditions. Thesuspension was centrifuged at 8000×g for 0.5 h and the supernatant wasremoved. The pellet was weighed and resuspended in fresh portion ofserum-free DMEM of a known weight. Known amount of doxorubicin andPluronic were dissolved in the suspension, which was allowed toequilibrate at 37° C. overnight, then snap-frozen and lyophilized. Thepowders were kept at −70° C. Before the transport experiments, a knownamount of powder was reconstituted in DMEM and was allowed toequilibrate at 37° C. overnight. The composition of the drug-containingsamples tested in the transport experiments is given in Table 3. TABLE 3Composition of the drug-containing donor media used in transepithelialtransport experiments in the apical or basolateral compartments. ^(a)Microgel Concentration of additive Microgel concentration, (beyonddoxorubicin), No. composition ^(b) μg/mL μg/mL ^(c) 1 None (Control) 0None 2 None 0 100 (Pluronic L61) 0 (Verapamil) 3 None 0 100 (PluronicL92) 0 (Verapamil) 4 L92-PAA-EGDMA 100 None 5 L92-PAA-EGDMA 100 100(Pluronic L61) 0 (Verapamil) 6 L92-PAA-EGDMA 100 100 (Pluronic L92) 0(Verapamil) 7 F127-PAA-EGDMA 100 0 8 F127-PAA-EGDMA 100 100 (PluronicL61) 0 (Verapamil) 9 F127-PAA-EGDMA 100 100 (Pluronic L92) 0 (Verapamil)10 None 0 0 (Pluronic) 9 (Verapamil) 11 L92-PAA-EGDMA 100 0 (Pluronic) 9(Verapamil) 12 F127-PAA-EGDMA 100 0 (Pluronic) 9 (Verapamil)^(a) Initial concentration of doxorubicin in the donor compartment was100 μg/mL in all experiments^(b) Degree of cross-linking, XL = 0.1 mol % throughout^(c) Concentrations of added Pluronic L61 and L92 were arbitrarilychosen to result in effective Pluronic concentrations just below CMC[40, 41]. Concentration of added Verapamil was 20 μM [27].

Cells were seeded at a density of (2-4)×10⁴ cells/cm² on top ofTranswell™ polycarbonate filters (pore size, 0.4 μm; diameter, 24.5 mm;growth area, 4.71 cm²) from Costar (Cambridge, Mass.). The cells weregrown for 3 weeks prior to the transport experiments andtrans-epithelial electrical resistance (TEER) was measured using aMillicell-ERS device (Millipore, Bedford, Mass.) equipped withrod-shaped electrodes. The TEER data were corrected for backgroundreadings contributed by the blank filter and culture medium. Typicalvalues of TEER were 800-850 Ωcm². Then the monolayers were rinsed twiceby serum-free DMEM and the transport experiment commenced by replacingthe medium at either the basolateral or the apical side of the monolayerwith 2.5 mL of the serum-free DMEM containing doxorubicin (with orwithout microgels, see Table 3). Simultaneously, the medium on the otherside was refreshed. The monolayers were incubated at 37° C. in 10% CO₂atmosphere. Samples of 200 μL were taken from each side intermittently,and the drug concentration was measured in the samples withdrawn fromthe side opposite to the side of the drug application. The volumeswithdrawn were immediately replaced with equal volumes of serum-freeDMEM pre-equilibrated at 37° C. Each experiment was repeated four times.The TEER values were measured after the completion of each transportexperiment and were shown to be equal to the initial TEER valuesobtained prior to the commencement of the experiment. The doxorubicinconcentration was assayed using a Shimadzu Model RF-5301 PCspectrofluorophotometer (λ_(excitation) 480 nm, λ_(emission) 580 nm).Additionally, the concentration of doxorubicin in the medium wasmeasured by HPLC using a Capcell Pak UG C18 (100×4.6 mm I.D., particlesize 5 μm) column and a Universal Guard Cartridge System (Phenomenex,Torrance, Calif.). The HPLC was a Hewlett-Packard 1090 system with anautosampler and a ZETALIF laser induced fluorescence detector (ESA,Inc., Chelmsford, Mass.). Water/acetonitrile (70/30, pH 4) was used asthe mobile phase (flow rate, 1 mL/min, injection volume, 5 μL), anddetection was carried out using an Argon Ion laser (488 nm, 10 mW).Surface tension in microgel suspensions was measured using the Wilhelmyplate method (Sigma 701 automatic tensiometer, KSV Instruments, Ltd.).Temperature was controlled to ±0.05 C using a circulating water bath.The platinum Wilhelmy plate was washed with acetone, rinsed in Milli-Qwater, and flamed until red-hot before each measurement.

The apparent permeability of the Caco-2 monolayer (P_(a), cm/s) wascalculated from the linearized time course of the doxorubicin fractionaltransport (dI₁t/dt in the fluorescence assay and dC_(t)/dt in the HPLCassay) normalized to the effective surface area of the filter (A=4.71cm²) and the initial fluorescence emission intensity reading (I_(o)) orinitial doxorubicin concentration (C_(o)) in the donor (basolateral orapical) compartment of volume V=2.5 cm³:$P_{a} = {{\frac{{\mathbb{d}I_{t}}/{\mathbb{d}t}}{{AI}_{0}}V} = {\frac{{\mathbb{d}C_{t}}/{\mathbb{d}t}}{{AC}_{0}}V}}$Excellent correlation was obtained between permeability valuesdetermined via fluorescence and HPLC assay (FIG. 14).

Transport Via Paracellular Route

The effect of microgel addition on paracellular permeability of theCaco-2 cell layers was estimated via permeability of mannitol, a neutralmolecule, which is absorbed exclusively by passive diffusion through theparacellular route [42].

To attenuate the effect of PAA on the transepithelial transport, theexperiments were conducted in calcium- and magnesium-free HBSS. Prior tothe commencement of the transport experiments, the culture DMEM wasreplaced with an equal volume of HBSS and the cells were allowed toequilibrate for 1 h. Donor suspensions (2.5 mL total) containing 0.1 or0.5 mg/mL microgels or Carbopol 934NF pre-equilibrated with ¹⁴C-mannitolin HBSS (initial specific activity of 0.2 μCi/mL) were used to replacethe HBSS on the apical side, and the experiment commenced after 10 minof initial equilibration. Permeability experiments were conducted at pH7.2, 37° C., 5% CO₂, and 90% relative humidity The TEER was measuredfollowing equilibration as described above for doxorubicin transport.TEER measurements were also performed during the experiment with 0.5mg/mL polymer loading in order to check the effect of polymers on theopening, if any, of the tight intercellular junctions at time intervalsof 0 (i.e. 10 min after adding the polymers), 30, 60, 90, 120, 150, 180,210, and 240 min. The samples withdrawn prior to the 30-min timeinterval were not included in the P_(m) calculation to ensuresteady-state kinetics. The withdrawn samples of ¹⁴C-mannitol were mixedwith 3 ml of MicroScint™ scintillation cocktail and the amount ofradioactive marker transported at each time interval was determinedusing a TopCount NXT scintillation counter (PerkinElmer Life Sciences,Boston, Mass.). For negative control, no polymer was applied to themonolayers. Samples of 200 μL were withdrawn from the basolateralchamber at predetermined time intervals of 0, 5, 15, 30, 60, 90, 120,180, 210, and 240 min and replaced with equal volumes of fresh HBSS.After completion of the transport studies, the polymers were removedcarefully and monolayers were rinsed with HBSS and the culture medium(DMEM) was applied on the monolayers. The monolayers were allowed toregenerate for 2 days at 37° C. in an atmosphere of 95% air and 5% CO₂at 90% relative humidity. TEER was monitored at 5, 6, 24, and 48 hduring the recovery period. Control transport experiments were alsoconducted across Transwell™ filters without Caco-2 cells to determinethe filter permeability (P_(filter)). The permeability of Caco-2 cellmonolayers (P_(m)) was estimated by correcting the effectivepermeability (P_(eff)) for filter permeability (P_(filter)) according tothe expression P_(eff) ⁻¹=P_(m) ⁻¹+P_(filter) ⁻¹.

Flow Cytometry

Intracellular accumulation of doxorubicin in Caco-2 cells viaP-gp-mediated efflux was studied by flow cytometry. Caco-2 cells(3×10⁵/cm²) were seeded into 24-well plates and incubated for 3 weeks asdescribed above. Cells were rinsed twice with phosphate-buffered saline(PBS) and pre-incubated for 30 min at 37° C. in 100 μg/mL PBS suspensionof microgels, Carbopol 934NF, or Pluronic L61 or L92 (pH 7.4). At theend of the 30-min incubation period, doxorubicin was added to theculture medium to result in its concentration of 1 μg/mL. Following 3-hincubation at 37° C., the cells were washed twice with ice-cold PBS.Then the cells were rinsed with 1 mM EDTA and 0.25% trypsin solution,collected into centrifuge tubes and centrifuged at 1000×g for 15 min,and finally resuspended in cold PBS. An aliquot of cells was kept on icefor analysis of dye or drug uptake.

Samples were analyzed on a FACScan flow cytometer (Becton DickinsonImmunocytometry Systems, San Jose, Calif.) equipped with a SpectraPhysics 15-mW argon laser (λ_(ex)=488 nm) and a red 585/42 band passfilter/FL2 fluorescence detector. All flow cytometric data were acquiredand analyzed with the CellQuest software (Becton Dickinson). Scatteringsignals were measured, collected, and corrected for, in the linear scalemode. The logarithmically amplified fluorescence emission intensity wasconverted to a linear scale and expressed in arbitrary units relative tothe control sample fluorescence intensity. The negative control wasperformed in drug-free medium to measure the cell auto-fluorescence. Thecontrol experiment was performed as described above, but withmicrogel-free PBS in the pre-incubation stage. At least 10⁴ cells wereanalyzed in each sample. Each experiment was repeated six times.

Colorimetric Cytotoxicity Assay

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assaycomprises cleavage of the tetrazolium salt to a dark blue product(formazan) by mitochondrial dehydrogenases in living but not in deadcells [43]. Caco-2 cells used for MTT assay were seeded onto 96 wellculture plates at seeding density of 5×10⁴ cells per well in DMEMculture medium. The cells were cultured in an atmosphere of 95% air and5% CO₂ at 37° C. and 90% humidity for 48 h. The culture medium wassubsequently replaced with HBSS and microgel suspension was added to thewells to result in a 0.5 mg/mL effective polymer concentration in eachwell. As negative control no polymer was added to the wells, and as aninternal reference, 0.5 mg/mL Carbopol in HBSS was applied to the cells.After adding the polymers, Caco-2 cells were further incubated at 37° C.for 4 h. The polymers were then removed, a 5 mg/ml MTT solution in PBSwas added to each well and the cells were incubated for another 4 h at37° C. The reaction product was then solubilized in dimethylsulfoxidebefore quantifying the color of the reaction product using an Emaxprecision microplate reader (Molecular Devices Co., Sunnyvale, Calif.)at 590 nm. In each MTT assay every microgel was tested in fivereplicates in microplate wells.

Statistical Analysis

All experiments were conducted at least in triplicate. The data wereanalyzed by Student's t-test at α=0.05. A one-tailed t-test (MicrosoftExcel®) (P<0.01) was used to identify significant differences betweenpermeability results with additives and in the control experiments.

Example Results

The apparent permeabilities of the Caco-2 cell layers to doxorubicinobtained in the transport experiments are collected in Table 4 infra. Asis seen, doxorubicin exhibited highly polarized transport, with theactive efflux exceeding the passive influx 4.6-fold in the case when noadditives were used. However, microgels and Pluronics (especiallyPluronic L92), as well as their combinations lowered the active effluxof doxorubicin from Caco-2 cells as much as 2.4-3.2-fold. Pluronic L61is known to be a potent doxorubicin efflux suppressor at concentrationsup to CMC [37, 40]. At concentrations above CMC, the effects ofPluronics generally plateau and then decay [34, 37, 40]. Surface tensionof 1100 μg/mL suspensions of L92-PAA-EGDMA and F127-PAA-EGDMA microgels(XL=0.1 mol %) at 37° C. (deionized water, pH adjusted to 7.4) wasmeasured to be 38.5 and 40.4 mN/m, respectively. That is, due to thenumerous Pluronic chains exposed to the aqueous environment, themicrogels exhibit surface activity comparable to the one of thehydrophobic Pluronic L61 or L92. Therefore, it can be hypothesized, withsome certainty, that Pluronic-PAA copolymers, which are surface-activeand solubilize lipids [14, 17, 44], can affect the membrane proteins ina fashion analogous to Pluronics. Verapamil, a known non-selectiveinhibitor of the P-gp, showed most dramatic effect on P_(a) ^(b→a)values (Table 4). Notably, this effect was not significantly neutralizedby the presence of anionic microgels, indicating that the apparent ionexchange kinetics of the organic base (Verapamil) between microgels andthe donor solutions were sufficiently rapid.

The passive influx of doxorubicin into Caco-2 cells was enhanced by allstudied additives, with the microgels exhibiting the most pronouncedeffects (up to 2.5-fold increase in P_(a) ^(a→b), Table 4). It has beenshown that absorption enhancers such as surfactants can act by improvingdrug absorption via paracellular (primarily hydrophilic drugs) as wellas transcellular (mostly lipophilic drugs) routes [45]. Combination ofenhanced transcellular passive influx and suppressed P-gp-mediatedactive influx leads to a significant accumulation of doxorubicin inCaco-2 cells and in MDR cancer cells when Pluronics are applied atconcentrations below their CMC [38, 39]. No significant enhancement ofthe paracellular drug absorption by the “unimeric” Pluronics (i.e. inthe absence of their micelles) have been reported [40, 46]. On the otherhand, surfactants are generally known to enhance paracellular transportby opening the tight junctions through an increase in the membrane poreradius, widening of the intercellular space, contraction ofcalmodulin-dependent actin microfilaments, or contraction of theperijunctional actomyosin ring [47-49]. In addition, the high capacityof poly(acrylic acid) to bind Ca²⁺ can deplete this ion from theextracellular cell medium and thus increase the paracellularpermeability of the epithelial cell layers [50, 51]. Indications existthat Pluronic-PAA can also bind Ca²⁺ in biological milieu [52].Therefore, it was important to address the question to what extent thedramatic increase in the net absorption by the Pluronic-PAA microgels(alone and in combination with Verapamil, Table 4) is due to theenhancement of the paracellular permeability and whether the microgelsare toxic to the cells. The flow cytometry study that was carried out toestimate the effects of the microgels on the intracellular accumulationof doxorubicin after 3-h incubation, showed significant enhancement ofthe doxorubicin uptake. That is, the enhancement factor was measured tobe 2.03±0.09 and 1.78±0.08 for the L92-PAA-EGDMA and F127-PAA-EGDMAmicrogels, respectively. In comparison, the enhancement factors forCarbopol, Pluronic L61, and L92 were measured to be 1.15±0.04,1.52±0.08, and 1.92±0.09, respectively. Herein, we define theenhancement factor as the ratio of the integrated fluorescence intensityin the experiment with microgel to the intensity in the controlexperiment, both corrected for natural cell fluorescence. Thus the flowcytometry results demonstrated significant enhancement of theintracellular uptake by both microgels and Pluronics alone and hinted atthe prevalence of the paracellular route. TABLE 4 Effect of microgels,Pluronic L61, Pluronic L92, and Verapamil on doxorubicin transport^(a)and absorption by Caco-2 cells. Treatment (Expt. No. in Table 1)P_(a)^(b → a) × 10⁶, cm/s P_(a)^(a → b) × 10⁶, cm/s Net absorption(secretion)^(b), % Control (1) 2.81 ± 0.03 0.61 ± 0.04 (360) PluronicL61 (2) 0.89 ± 0.04 1.12 ± 0.03 −21 Pluronic L92 (3) 0.68 ± 0.04 1.17 ±0.03 −42 L92-PAA-EGDMA (4) 1.12 ± 0.03 1.55 ± 0.03 −28 L92-PAA-EGDMA +Pluronic L61 (5) 0.95 ± 0.04 1.46 ± 0.03 −35 L92-PAA-EGDMA + PluronicL92 (6) 0.59 ± 0.04 1.26 ± 0.04 −53 F127-PAA-EGDMA (7) 1.12 ± 0.04 1.32± 0.04 −15 F127-PAA-EGDMA + Pluronic L61 (8) 1.16 ± 0.03 1.44 ± 0.04 −19F127-PAA-EGDMA + Pluronic L92 (9) 0.57 ± 0.03 1.23 ± 0.04 −54 Verapamil(10) 0.45 ± 0.05 0.88 ± 0.04 −49 L92-PAA-EGDMA + verapamil (11) 0.49 ±0.05 1.39 ± 0.05 −65 F127-PAA-EGDMA + verapamil (12) 0.54 ± 0.05 1.29 ±0.05 −58 ^(a)Apparent permeabilities with P ≦ 0.05${{\,^{b}{Calculated}}\quad{using}\quad{the}\quad{{formula}:\quad{{Net}\quad{effect}}}} = {100 \times \frac{P_{a}^{b\rightarrow a} - P_{a}^{a\rightarrow b}}{P_{a}^{a\rightarrow b}}}$

Integrity of Caco-2 cell monolayers and opening of tight junctions canbe assessed by measuring the flux of small hydrophilic radiolabeledmolecules such as ¹⁴C-mannitol across the monolayers, as well as by TEER[53]. Transport of ¹⁴C-mannitol across Caco-2 monolayers was assessed inthe presence of microgels as well as benign polymer widely used in oralapplications (Carbopol 934NF), using HBSS as a negative control. Asmeasured by radioactivity count, total cumulative transport of¹⁴C-mannitol was relatively minor in all instances and did not exceed3.5% of initial concentration in the donor compartment (FIG. 15).

Using the relative release kinetics, the effective permeability(P_(eff)) of monolayers was calculated as described in Experimentalsection (Table 5). As is seen, at concentrations of 0.1 mg/mL (as inTable 4) either Carbopol or microgels did not significantly enhance theP_(eff). At concentrations of 0.5 mg/mL, the microgels and Carbopolsignificantly increased the P_(eff) as compared to the negative control,but the differences between the microgels and Carbopol wereinsignificant. This is an important result indicating that although ourmicrogels do result in the enhancement of the intracellular doxorubicintransport (Table 4), they do not provoke any dramatic changes in theparacellular permeability. The microgels compared favorably withCarbopol (lightly cross-linked poly(acrylic acid)), which is an industrystandard in formulations that require adhesion to gastrointestinaltissues [54, 55]. TABLE 5 Effective permeability (P_(eff)) of Caco-2cell monolayers (mean ± S.D. of 3-4 experiments) for ¹⁴C-mannitol.Polymer concentration, P_(eff) ×10⁷, Permeability Sample/Additive mg/mLcm/s ratio ^(a) Control 0 2.74 ± 0.44 1.0 Carbopol 0.1 3.36 ± 0.75 1.20.5 5.05 ± 1.45 1.8 L92-PAA-EGDMA 0.1 3.39 ± 0.92 1.2 0.5 3.89 ± 0.831.4 F127-PAA-EGDMA 0.1 4.43 ± 1.39 1.6 0.5 7.58 ± 1.26 2.8^(a) Relative to the Control experiment (P ≦ 0.05).

The observed tendency of added polymers (at 0.5 mg/mL level) to increasepermeability of Caco-2 layers might be an indication that the polymersaffect the integrity of cell membrane. Therefore, the reversibility ofthis effect is an important issue when screening these polymers aspenetration enhancers. Herein, it was observed that after removing thepolymers from the monolayers, TEER values completely recovered toinitial values within 2 days, indicating that the effects, if any, ofmicrogels on tight junctions are fully reversible (FIG. 16). It shouldbe noted that at 0.1 mg/mL level, where microgels exhibited significanteffects on the net absorption of doxorubicin (Table 4), the TEER was notsignificantly affected, which is an evidence that the transport acrossthe Caco-2 monolayers in the presence of microgels is dominated by thetranscellular, and not paracellular, pathway.

The MTT assay showed no toxic effects caused by mucosal application ofthe microgels in comparison with the negative control: 99±12, 97±11 and97±15% of the cells were viable after application of microgelsL92-PAA-EGDMA, F127-PAA-EGDMA, and Carbopol, respectively. Using the MTTassay, the polymer-treated cells were able to metabolize themitochondrial substrate MTT by conversion into formazan crystals. Thismetabolic activity of cells is an appropriate technique for assessingthe number of viable cells, since damaged or dead cells are devoid ofany mitochondrial dehydrogenase activity [56]. Thus the Caco-2 cellmonolayers appeared to be viable after 4 h application of microgels andno damage was observed at the intracellular level. Overall, the effectof microgels on mitochondrial dehydrogenase activity revealed theirbenign nature.

By inhibiting the P-gp-mediated doxorubicin efflux from the cells andenhancing the passive influx, the lightly cross-linked Pluronic-PAAmicrogels enhance the overall absorption of the drug by the cells.Notably, this effect is more pronounced that with a known penetrationenhancer, Pluronic L61, and is comparable to the other relativelyhydrophobic copolymer, Pluronic L92. Microgels exhibit synergism withVerapamil, a non-selective inhibitor of the P-gp. Judging by¹⁴C-mannitol permeability, the microgels do not damage cells, so that nomeaningful enhancement of the paracellular transport is observed. Anyeffect of microgels on trans-epithelial electrical resistance appears tobe fully reversible. Notably, materials comprising slightly cross-linkedpoly(acrylic acid) have demonstrated no systemic absorption ingastrointestinal transit experiments both in vitro and in vivo [57, 58].

Given the benign, non-irritating nature and mucoadhesive properties ofthe Pluronic-PAA microgels, for example, their application in theformulations for oral delivery of anticancer drugs is valuable indeed.

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All publications and patents mentioned in this specification are hereinincorporated by reference. Various modifications and variations of thedescribed composition of matter, methods of manufacture and methods ofuse of the invention will be apparent to those skilled in the artwithout departing from the scope and spirit of the invention. Althoughthe invention has been described in connection with specific preferredembodiments, it should be understood that the invention as claimedshould not be unduly limited to such specific embodiments. Indeed,various modifications of the described modes for carrying out theinvention which are obvious to those skilled in polymer chemistry andformulation are intended to be within the scope of the following claims.

1-3. (canceled)
 4. A responsive microgel which comprises: an ionizablenetwork of covalently cross-linked homopolymeric ionizable monomerswherein the ionizable network is covalently attached to an amphiphiliccopolymer to form a plurality of ‘dangling chains’ and wherein the‘dangling chains’ of amphiphilic copolymer form immobile aggregates inaqueous solution; and at least one therapeutic entity.
 5. A responsivemicrogel according to claim 4 which comprises a cationic therapeuticentity.
 6. A responsive microgel according to claim 4 which comprises ahydrophobic therapeutic entity.
 7. A responsive microgel according toclaim 4 which comprises an amphiphilic therapeutic entity.
 8. Aresponsive microgel according to claim 4 which delivers a substantiallylinear and sustained release of a hydrophobic or amphiphilic therapeuticentity under physiological conditions. 9.-16. (canceled)
 17. A method ofadministering at least one therapeutic entity to a patient comprisingadministering a responsive microgel according to claim
 4. 18. A methodof administering at least one therapeutic entity to a patient accordingto claim 17 comprising orally administering a responsive microgel.
 19. Amethod of administering at least one therapeutic entity to a patientaccording to claim 17 selected from the group consisting of (hydrophobicentities, cationic entities, and amphiphilic entities).
 20. A method ofadministering at least one therapeutic entity to a patient according toclaim 17 selected from the group consisting of (a steroidalantiandrogen, a non steroidal antiandrogen, an estrogen,diethylstilbestrol, a conjugated estrogen, a selective estrogen receptormodulator (SERM), a taxane, a LHRH analog, substrates of ABCtransporters such as P-glycoprotein; MRP1-MRP9; ABC half-transporterssuch as BCRP and other transporters that are involved into a limiteddrug transport across small intestinal epythlium, cerebral endotheliumand other barrier tissues in the body, as well as substrates ofmetabolic enzyme isoforms without limitation, cytochrome P-450;esterase; epoxide hydrolase; alcohol dehydrogenase; aldehydedehydroganase; dihydropyrimidine dehydroganase; NADPH-quinoneoxidoreductase; uridine 5′-triphosphat glucoronosyltransferase;sulfotransferase; glutatione S-transferase; N-acetiltransferase;histamine methyltransferase; catechol-o-methyl transferase; thiopurinemethyltransferase. This group of therapeutic agents include withoutlimitation doxorubicin and other anthracyclines, mitoxantrone, mitomycinC, methotrexate, paclitaxel, docetaxel and other taxanes, topotecan andother camptotecines, cysplatin, carboplatin, oxaliplatin and otherplatinum complexes; megesterol acetate and other steroids; carvediloland other beta-blocking agents; azidothymidine, fludarabine and othernucleoside containing agents in their dephospho, mono-, di- andtri-phosphorylated forms; vinblastine, vincristine and other vinkaalkaloids; etoposide and other podophilotoxins).